Promoters from Brassica napus for seed specific gene expression

ABSTRACT

The present invention is concerned with means and methods for allowing tissue specific and, in particular, seed specific expression of genes. The present invention, accordingly, relates to a polynucleotide comprising an expression control sequence which allows seed specific expression of a nucleic acid of interest being operatively linked thereto. Moreover, the present invention contemplates vectors, host cells, non-human transgenic organisms comprising the aforementioned polynucleotide as well as methods and uses of such a polynucleotide.

RELATED APPLICATIONS

This application is a national stage application (under 35 U.S.C. §371) of PCT/EP2009/058138, filed Jun. 30, 2009, which claims benefit of European application 08159440.0, filed Jul. 1, 2008.

SUBMISSION OF SEQUENCE LISTING

The Sequence Listing associated with this application is filed in electronic format via EFS-Web and hereby incorporated by reference into the specification in its entirety. The name of the text file containing the Sequence Listing is Sequence_Listing_(—)17418_(—)00062. The size of the text file is 416 KB, and the text file was created on Dec. 28, 2010.

FIELD OF THE INVENTION

The present invention is concerned with means and methods for allowing tissue specific and, in particular, seed specific expression of genes. The present invention, accordingly, relates to a polynucleotide comprising an expression control sequence which allows seed specific expression of a nucleic acid of interest being operatively linked thereto. Moreover, the present invention contemplates vectors, host cells, non-human transgenic organisms comprising the aforementioned polynucleotide as well as methods and uses of such a polynucleotide.

BACKGROUND OF THE INVENTION

In the field of “green” (agricultural) biotechnology, plants are genetically manipulated in order to confer beneficial traits. These beneficial traits may be yield increase, tolerance increase, reduced dependency on fertilizers, herbicidal, pesticidal- or fungicidal-resistance, or the capability of producing chemical specialties such as nutrients, drugs, oils for food and petrochemistry etc.

In many cases, it is required to express a heterologous gene in the genetically modified plants at a rather specific location in order to obtain a plant exhibiting the desired beneficial trait. One major location for gene expression is the plant seed. In the seeds, many important synthesis pathways, e.g., in fatty acid synthesis, take place. Accordingly, expression of heterologous genes in seeds allow for the manipulation of fatty acid synthesis pathways and, thus, for the provision of various fatty acid derivatives and lipid-based compounds.

However, for many heterologous genes, a seed specific expression will be required. Promoters which allow for a seed specific expression are known in the art. Such promoters include the oilseed rape napin promoter (U.S. Pat. No. 5,608,152), the Vicia faba USP promoter (Baeumlein et al., Mol Gen Genet, 1991, 225 (3):459-67), the Arabidopsis oleosin promoter (WO 98/45461), the Phaseolus vulgaris phaseolin promoter (U.S. Pat. No. 5,504,200), the Brassica Bce4 promoter (WO 91/13980) or the legumine B4 promoter (LeB4; Baeumlein et al., 1992, Plant Journal, 2 (2):233-9), and promoters which bring about the seed-specific expression in monocotyledonous plants such as maize, barley, wheat, rye, rice and the like. Suitable noteworthy promoters are the barley Ipt2 or Ipt1 gene promoter (WO 95/15389 and WO 95/23230) or the promoters from the barley hordein gene, the rice glutelin gene, the rice oryzin gene, the rice prolamine gene, the wheat gliadine gene, the wheat glutelin gene, the maize zeine gene, the oat glutelin gene, the sorghum kasirin gene or the rye secalin gene, which are described in WO 99/16890.

However, there is a clear need for further expression control sequences such as promoters and terminators which allow for a reliable and efficient control of expression of foreign nucleic acids in seeds.

The technical problem underlying this invention can be seen as the provision of means and methods complying with the aforementioned needs. The technical problem is solved by the embodiments characterized in the claims and herein below.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows a developmental expression analysis. Tissue types and developmental stages are given as listed in table 10. Samples 10, 11, 12 were pooled (assigned as 10), as well as samples 13, 14, 15 (assigned as 13).

FIG. 2 shows the 18:3n-6 (GLA) content of seeds-oil of seeds harvested from transgenic plants harboring the T-DNA from vectors described in example 3. Shown are data from T1 seeds and T2 seeds as indicated on top of the figure. Measurements on T1 on seeds are on individual single seeds; measurements on T2 seeds are on seed batches. The black line indicates the minimal and the maximal observation, the box reaches from the 25% quartil to the 75 quartil; the median is indicated as black line within the box. The number of individual measurements is indicated as number above each box.

FIG. 3 shows the 18:3n-6 (GLA) and 18:4n-3 (SDA) content of seeds-oil of seeds harvested from transgenic Arabidopsis plants harboring the T-DNA from vectors described in example 3.

FIG. 4 illustrates for the different promoters the different ratios of the omega-3 fatty acid SDA to the omega-6 fatty acid GLA.

DETAILED DESCRIPTION OF THE INVENTION

Accordingly, the present invention relates to a polynucleotide comprising an expression control sequence which allows seed specific expression of a nucleic acid of interest being operatively linked thereto, said expression control sequence being selected from the group consisting of:

-   -   (a) an expression control sequence having a nucleic acid         sequence as shown in any one of SEQ ID NOs: 1, 6, 9, 14, 16, 22,         25, 70, 77, 85, 95, 103, 111, 119, 124 or 131;     -   (b) an expression control sequence having a nucleic acid         sequence which is at least 80% identical to a nucleic acid         sequence shown in any one of SEQ ID NOs: 1, 6, 9, 14, 16, 22,         25, 70, 77, 85, 95, 103, 111, 119, 124 or 131;     -   (c) an expression control sequence having a nucleic acid         sequence which hybridizes under stringent conditions to a a         nucleic acid sequence as shown in any one of SEQ ID NOs: 1, 6,         9, 14, 16, 22, 25, 70, 77, 85, 95, 103, 111, 119, 124 or 131;     -   (d) an expression control sequence having a nucleic acid         sequence which hybridizes to a nucleic acid sequences located         upstream of an open reading frame sequence shown in any one of         SEQ ID NOs: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 or         125;     -   (e) an expression control sequence having a nucleic acid         sequence which hybridizes to a nucleic acid sequences located         upstream of an open reading frame sequence being at least 80%         identical to an open reading frame sequence as shown in any one         of SEQ ID NOs: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112         or 125;     -   (f) an expression control sequence obtainable by 5′ genome         walking or by thermal asymmetric interlaced polymerase chain         reaction (TAIL-PCR) on genomic DNA from the first exon of an         open reading frame sequence as shown in any one of SEQ ID NOs:         5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 or 125; and     -   (g) an expression control sequence obtainable by 5′ genome         walking or TAIL PCR on genomic DNA from the first exon of an         open reading frame sequence being at least 80% identical to an         open reading frame as shown in any one of SEQ ID NOs: 5, 13, 20,         21, 27, 28, 71, 78, 86, 96, 104, 112 or 125.

The term “polynucleotide” as used herein refers to a linear or circular nucleic acid molecule. It encompasses DNA as well as RNA molecules. The polynucleotide of the present invention is characterized in that it shall comprise an expression control sequence as defined elsewhere in this specification. In addition to the expression control sequence, the polynucleotide of the present invention, preferably, further comprises at least one nucleic acid of interest being operatively linked to the expression control sequence and/or a termination sequence or transcription. Thus, the polynucleotide of the present invention, preferably, comprises an expression cassette for the expression of at least one nucleic acid of interest. Alternatively, the polynucleotide may comprise in addition to the said expression control sequence a multiple cloning site and/or a termination sequence for transcription. In such a case, the multiple cloning site is, preferably, arranged in a manner as to allow for operative linkage of a nucleic acid to be introduced in the multiple cloning site with the expression control sequence. In addition to the aforementioned components, the polynucleotide of the present invention, preferably, could comprise components required for homologous recombination, i.e. flanking genomic sequences from a target locus. However, also preferably, the polynucleotide of the present invention can essentially consist of the said expression control sequence.

The term “expression control sequence” as used herein refers to a nucleic acid which is capable of governing the expression of another nucleic acid operatively linked thereto, e.g. a nucleic acid of interest referred to elsewhere in this specification in detail. An expression control sequence as referred to in accordance with the present invention, preferably, comprises sequence motifs which are recognized and bound by polypeptides, i.e. transcription factors. The said transcription factors shall upon binding recruit RNA polymerases, preferably, RNA polymerase I, II or III, more preferably, RNA polymerase II or III, and most preferably, RNA polymerase II. Thereby the expression of a nucleic acid operatively linked to the expression control sequence will be initiated. It is to be understood that dependent on the type of nucleic acid to be expressed, i.e. the nucleic acid of interest, expression as meant herein may comprise transcription of RNA polynucleotides from the nucleic acid sequence (as suitable for, e.g., anti-sense approaches or RNAi approaches) or may comprises transcription of RNA polynucleotides followed by translation of the said RNA polynucleotides into polypeptides (as suitable for, e.g., gene expression and recombinant polypeptide production approaches). In order to govern expression of a nucleic acid, the expression control sequence may be located immediately adjacent to the nucleic acid to be expressed, i.e. physically linked to the said nucleic acid at its 5″end. Alternatively, it may be located in physical proximity. In the latter case, however, the sequence must be located so as to allow functional interaction with the nucleic acid to be expressed. An expression control sequence referred to herein, preferably, comprises between 200 and 5,000 nucleotides in length. More preferably, it comprises between 500 and 2,500 nucleotides and, more preferably, at least 1,000 nucleotides. As mentioned before, an expression control sequence, preferably, comprises a plurality of sequence motifs which are required for transcription factor binding or for conferring a certain structure to the polynucletide comprising the expression control sequence. Sequence motifs are also sometimes referred to as cis-regulatory elements and, as meant herein, include promoter elements as well as enhancer elements. Preferred expression control sequences to be included into a polynucleotide of the present invention have a nucleic acid sequence as shown in any one of SEQ ID NOs: 1, 6, 9, 14, 16, 22, 25, 70, 77, 85, 95, 103, 111, 119, 124 and 131.

Further preferably, an expression control sequence comprised by a polynucleotide of the present invention has a nucleic acid sequence which hybridizes to a nucleic acid sequences located upstream of an open reading frame sequence shown in any one of SEQ ID NOs: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 and 125, i.e. is a variant expression control sequence. It will be understood that expression control sequences may slightly differ in its sequences due to allelic variations. Accordingly, the present invention also contemplates an expression control sequence which can be derived from an open reading frame as shown in any one of SEQ ID NOs: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 and 125. Said expression control sequences are capable of hybridizing, preferably under stringent conditions, to the upstream sequences of the open reading frames shown in any one of SEQ ID NOs. 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 and 125, i.e. the expression control sequences shown in any one of SEQ ID NOs.: 1, 6, 9, 14, 16, 22, 25, 70, 77, 85, 95, 103, 111, 119, 124 and 131. Stringent hybridization conditions as meant herein are, preferably, hybridization conditions in 6× sodium chloride/sodium citrate (=SSC) at approximately 45° C., followed by one or more wash steps in 0.2×SSC, 0.1% SDS at 53 to 65° C., preferably at 55° C., 56° C., 57° C., 58° C., 59° C., 60° C., 61° C., 62° C., 63° C., 64° C. or 65° C. The skilled worker knows that these hybridization conditions differ depending on the type of nucleic acid and, for example when organic solvents are present, with regard to the temperature and concentration of the buffer. For example, under “standard hybridization conditions” the temperature differs depending on the type of nucleic acid between 42° C. and 58° C. in aqueous buffer with a concentration of 0.1 to 5×SSC (pH 7.2). If organic solvent is present in the abovementioned buffer, for example 50% formamide, the temperature under standard conditions is approximately 42° C. The hybridization conditions for DNA:DNA hybrids are preferably for example 0.1×SSC and 20° C. to 45° C., preferably between 30° C. and 45° C. The hybridization conditions for DNA:RNA hybrids are preferably, for example, 0.1×SSC and 30° C. to 55° C., preferably between 45° C. and 55° C. The abovementioned hybridization temperatures are determined for example for a nucleic acid with approximately 100 bp (=base pairs) in length and a G+C content of 50% in the absence of formamide. Such hybridizing expression control sequences are, more preferably, at least 70%, at least 80%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94% at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the expression control sequences as shown in any one of SEQ ID NOs.: 1, 6, 9, 14, 16, 22, 25, 70, 77, 85, 95, 103, 111, 119, 124 and 131. The percent identity values are, preferably, calculated over the entire nucleic acid sequence region. A series of programs based on a variety of algorithms is available to the skilled worker for comparing different sequences. In this context, the algorithms of Needleman and Wunsch or Smith and Waterman give particularly reliable results. To carry out the sequence alignments, the program PileUp (J. Mol. Evolution., 25, 351-360, 1987, Higgins 1989, CABIOS, 5: 151-153) or the programs Gap and BestFit (Needleman 1970 J. Mol. Biol. 48; 443-453 and Smith 1981, Adv. Appl. Math. 2; 482-489), which are part of the GCG software packet (Genetics Computer Group, 575 Science Drive, Madison, Wis., USA 53711 version 1991), are to be used. The sequence identity values recited above in percent (%) are to be determined, preferably, using the program GAP over the entire sequence region with the following settings: Gap Weight: 50, Length Weight: 3, Average Match: 10.000 and Average Mismatch: 0.000, which, unless otherwise specified, shall always be used as standard settings for sequence alignments.

Moreover, expression control sequences which allow for seed specific expression can not only be found upstream of the aforementioned open reading frames having a nucleic acid sequence as shown in any one of SEQ ID NOs. 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 and 125. Rather, expression control sequences which allow for seed specific expression can also be found upstream of orthologous, paralogous or homologous genes (i.e. open reading frames). Thus, also preferably, an variant expression control sequence comprised by a polynucleotide of the present invention has a nucleic acid sequence which hybridizes to a nucleic acid sequences located upstream of an open reading frame sequence being at least 70%, more preferably, at least 80%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94% at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to a sequence as shown in any one of SEQ ID NOs: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 and 125. The said variant open reading shall encode a polypeptide having the biological activity of the corresponding polypeptide being encoded by the open reading frame shown in any one of SEQ ID NOs.: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 and 125. In this context it should be mentioned that the open reading frame shown in SEQ ID NO: 5 encodes a polypeptide showing similarity to gibberlin responsive proteins, the open reading frame shown in SEQ ID NO: 13 encodes a polypeptide belonging to the pectinesterase family, the open reading frame shown in SEQ ID NO: 20 encodes “sinapoyl choline transferase 1” (SCT1), and the open reading frames shown in SEQ ID NO: 21 encodes “sinapoyl choline transferase 2” (SCT2). These biological activities can be determined by those skilled in the art without further ado. The open reading frames shown in SEQ ID NO: 27 and 28 encode polypeptides showing homology to seed proteins with yet unknown functions.

TABLE 1 Protein function of genes identified to be seed specifically expressed: SEQ ID Protein function 5 putative gibberelin responsive protein 13 putative pectinesterase 20 Sinapoyl choline transferase 21 Sinapoyl choline transferase 27 Seed protein 28 Seed protein 72 Seed protein 80 CRU4 subunit of Cruciferin (seed storage protein) 88 Myrosinase 98 Seed protein 106 serine proteinase inhibitor 114 Transcription factor involved in embryonic development 121 Glutathione S-transferase 133 Seed protein

Also preferably, a variant expression control sequence comprised by a polynucleotide of the present invention is (i) obtainable by 5′ genome walking or TAIL PCR from an open reading frame sequence as shown in any one of SEQ ID NOs: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 and 125 or (ii) obtainable by 5′ genome walking or TAIL PCR from a open reading frame sequence being at least 80% identical to an open reading frame as shown in any one of SEQ ID NOs: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 and 125. Variant expression control sequences are obtainable without further ado by the genome walking technology or by thermal asymmetric interlaced polymerase chain reaction (TAIL-PCR) which can be carried out as described in the accompanying Examples by using, e.g., commercially available kits.

Variant expression control sequences referred to in this specification for the expression control sequence shown in SEQ ID NO: 1, preferably, comprise at least 80, at least 90, at least 100, at least 110, at least 120, at least 130, at least 140 or all of the sequence motifs recited in Table 1. Variant expression control sequences referred to in this specification for the expression control sequence shown in SEQ ID NO: 6, preferably, comprise at least 80, at least 90, at least 100, at least 110 or all of the sequence motifs recited in Table 2. Variant expression control sequences referred to in this specification for the expression control sequence shown in SEQ ID NO: 9, preferably, comprise at least 40, at least 50, at least 60 or all of the sequence motifs recited in Table 3. Variant expression control sequences referred to in this specification for the expression control sequence shown in SEQ ID NO: 14, preferably, comprise at least 50, at least 60, at least 70, at least 80, at least 90 or all of the sequence motifs recited in Table 4. Variant expression control sequences referred to in this specification for the expression control sequence shown in SEQ ID NO: 16, preferably, comprise at least 50, at least 60, at least 70, at least 80, at least 90 or all of the sequence motifs recited in Table 5. Variant expression control sequences referred to in this specification for the expression control sequence shown in SEQ ID NO: 22, preferably, comprise at least 80, at least 90, at least 100, at least 110, at least 120, at least 130 or all of the sequence motifs recited in Table 6. Variant expression control sequences referred to in this specification for the expression control sequence shown in SEQ ID NO: 25, preferably, comprise at least 80, at least 100, at least 120, at least 130, at least 140, at least 150 or all of the sequence motifs recited in Table 7.

Variant expression control sequences referred to in this specification also, preferably, comprise at least the cis-regulatory elements referred to in Table 8, below. Even more preferably, the variant regulatory expression control sequences comprise said elements with the same frequency and distribution as referred to in Table 9 for the individual regulatory sequences.

The term “seed specific” as used herein means that a nucleic acid of interest being operatively linked to the expression control sequence referred to herein will be predominantly expressed in seeds when present in a plant. A predominant expression as meant herein is characterized by a statistically significantly higher amount of detectable transcription in the seeds with respect to other plant tissues. A statistically significant higher amount of transcription is, preferably, an amount being at least two-fold, three-fold, four-fold, five-fold, ten-fold, hundred-fold, five hundred-fold or thousand-fold the amount found in at least one of the other tissues with detectable transcription. Alternatively, it is an expression in seeds whereby the amount of transcription in non-seed tissues is less than 1%, 2%, 3%, 4% or most preferably 5% of the overall (whole plant) amount of expression. The amount of transcription directly correlates to the amount of transcripts (i.e. RNA) or polypeptides encoded by the transcripts present in a cell or tissue. Suitable techniques for measuring transcription either based on RNA or polypeptides are well known in the art. Seed specific alternatively and, preferably in addition to the above, means that the expression is restricted or almost restricted to seeds, i.e. there is essentially no detectable transcription in other tissues. Almost restricted as meant herein means that unspecific expression is detectable in less than ten, less than five, less than four, less than three, less than two or one other tissue(s). Seed specific expression as used herein includes expression in seed cells or their precursors, such as cells of the endosperm and of the developing embryo.

An expression control sequences can be tested for seed specific expression by determining the expression pattern of a nucleic acid of interest, e.g., a nucleic acid encoding a reporter protein, such as GFP, in a transgenic plant. Transgenic plants can be generated by techniques well known to the person skilled in the art and as discussed elsewhere in this specification. The aforementioned amounts or expression pattern are, preferably, determined by Northern Blot or in situ hybridization techniques as described in WO 02/102970 in Brassica napus plants, more preferably, at 20, 25, 30, 35 or 40 days after flowering. Preferred expression pattern for the expression control sequences according to the present invention are shown in the Figure or described in the accompanying Examples, below.

The term “nucleic acid of interest” refers to a nucleic acid which shall be expressed under the control of the expression control sequence referred to herein. Preferably, a nucleic acid of interest encodes a polypeptide the presence of which is desired in a cell or non-human organism as referred to herein and, in particular, in a plant seed. Such a polypeptide may be an enzyme which is required for the synthesis of seed storage compounds or may be a seed storage protein. It is to be understood that if the nucleic acid of interest encodes a polypeptide, transcription of the nucleic acid in RNA and translation of the transcribed RNA into the polypeptide may be required. A nucleic acid of interest, also preferably, includes biologically active RNA molecules and, more preferably, antisense RNAs, ribozymes, micro RNAs or siRNAs. Said biologically active RNA molecules can be used to modify the amount of a target polypeptide present in a cell or non-human organism. For example, an undesired enzymatic activity in a seed can be reduced due to the seed specific expression of an antisense RNAs, ribozymes, micro RNAs or siRNAs. The underlying biological principles of action of the aforementioned biologically active RNA molecules are well known in the art. Moreover, the person skilled in the art is well aware of how to obtain nucleic acids which encode such biologically active RNA molecules. It is to be understood that the biologically active RNA molecules may be directly obtained by transcription of the nucleic acid of interest, i.e. without translation into a polypeptide. It is to be understood that the expression control sequence may also govern the expression of more than one nucleic acid of interest, i.e. at least one, at least two, at least three, at least four, at least five etc. nucleic acids of interest.

The term “operatively linked” as used herein means that the expression control sequence of the present invention and a nucleic acid of interest, are linked so that the expression can be governed by the said expression control sequence, i.e. the expression control sequence shall be functionally linked to said nucleic acid sequence to be expressed. Accordingly, the expression control sequence and, the nucleic acid sequence to be expressed may be physically linked to each other, e.g., by inserting the expression control sequence at the 5″end of the nucleic acid sequence to be expressed. Alternatively, the expression control sequence and the nucleic acid to be expressed may be merely in physical proximity so that the expression control sequence is capable of governing the expression of at least one nucleic acid sequence of interest. The expression control sequence and the nucleic acid to be expressed are, preferably, separated by not more than 500 bp, 300 bp, 100 bp, 80 bp, 60 bp, 40 bp, 20 bp, 10 bp or 5 bp.

Advantageously, it has been found in the studies underlying the present invention that seed specific expression of a nucleic acid of interest can be achieved by expressing said nucleic acid of interest under the control of an expression control sequence from Brassica napus or a variant expression control sequence as specified above. The expression control sequences provided by the present invention allow for a reliable and highly specific expression of nucleic acids of interest. Thanks to the present invention, it is possible to (i) specifically manipulate biochemical processes in seeds, e.g., by expressing heterologous enzymes or biologically active RNAs, or (ii) to produce heterologous proteins in seeds. In principle, the present invention contemplates the use of the polynucleotide, the vector, the host cell or the non-human transgenic organism for the expression of a nucleic acid of interest. Preferably, the envisaged expression is seed specific. More preferably, the nucleic acid of interest to be used in the various embodiments of the present invention encodes a seed storage protein or is involved in the modulation of seed storage compounds.

As used herein, seed storage compounds include fatty acids and triacylglycerides which have a multiplicity of applications in the food industry, in animal nutrition, in cosmetics and the pharmacological sector. Depending on whether they are free saturated or unsaturated fatty acids or else triacylglycerides with an elevated content of saturated or unsaturated fatty acids, they are suitable for various different applications. More preferably, the polynucleotide of the present invention comprising the expression control sequence referred to above is applied for the manufacture of polyunsaturated fatty acids (PUFAs). For the manufacture of PUFAs in seeds, the activity of enzymes involved in their synthesis, in particular, elongases and desaturases, needs to be modulated. This will be achieved by seed specific expression of the nucleic acids of interest encoding the aforementioned enzymes or by seed specific expression of antisense, ribozyme, RNAi molecules which downregulate the activity of the enzymes by interfering with their protein synthesis. PUFAs are seed storage compounds which can be isolated by a subsequently applied purification process using the aforementioned seeds.

Particularly preferred PUFAs in accordance with the present invention are polyunsaturated long-chain ω-3-fatty acids such as eicosapentaenoic acid (=EPA, C20:5^(Δ5,8,11,14,17)), ω-3 eicostetraenic acid (=ETA, C20:4^(Δ8,11,14,17)), arachidonic acid (=ARA C20:4^(Δ5,8,11,14)) or docosahexaenoic acid (=DHA, C22:6^(Δ4,7,10,13,16,19)). They are important components of human nutrition owing to their various roles in health aspects, including the development of the child brain, the functionality of the eyes, the synthesis of hormones and other signal substances, and the prevention of cardiovascular disorders, cancer and diabetes (Poulos, A Lipids 30:1-14, 14^(Δ8,11,14,17)995; Horrocks, L A and Yeo Y K Pharmacol Res 40:211-225, 1999). There is, therefore, a need for the production of polyunsaturated long-chain fatty acids.

Particular preferred enzymes involved in the synthesis of PUFAs are disclosed in WO 91/13972 (Δ9-desaturase), WO 93/11245 (Δ15-desaturase), WO 94/11516 (Δ12-desaturase), EP A 0 550 162, WO 94/18337, WO 97/30582, WO 97/21340, WO 95/18222, EP A 0 794 250, Stukey et al., J. Biol. Chem., 265, 1990: 20144-20149, Wada et al., Nature 347, 1990: 200-203 or Huang et al., Lipids 34, 1999: 649-659. Δ6-Desaturases are described in WO 93/06712, U.S. Pat. No. 5,614,393, U.S. Pat. No. 5,614,393, WO 96/21022, WO 00/21557 and WO 99/27111, and also the application for the production in transgenic organisms is described in WO 98/46763, WO 98/46764 and WO 98/46765. Here, the expression of various desaturases is also described and claimed in WO 99/64616 or WO 98/46776, as is the formation of polyunsaturated fatty acids. As regards the expression efficacy of desaturases and its effect on the formation of polyunsaturated fatty acids, it must be noted that the expression of a single desaturase as described to date has only resulted in low contents of unsaturated fatty acids/lipids such as, for example, γ-linolenic acid and stearidonic acid. Furthermore, mixtures of ω-3- and ω-6-fatty acids are usually obtained.

Furthermore, the present invention relates to a polynucleotide comprising an expression termination sequence which allows for termination of transcription of a nucleic acid of interest being operatively linked thereto, said expression termination sequence being selected from the group consisting of:

-   -   (a) a expression termination sequence having a nucleic acid         sequence as shown in any one of SEQ ID NOs: 2, 7, 10, 15, 17,         23, 71, 78, 86, 96, 104, 112, or 125;     -   (b) a expression termination sequence having a nucleic acid         sequence which is at least 80% identical to a nucleic acid         sequence as shown in any one of SEQ ID NOs: which hybridizes         under stringent conditions to a nucleic acid sequence as shown         in any one of SEQ ID NOs: 2, 7, 10, 15, 17, 23, 71, 78, 86, 96,         104, 112, or 125;     -   (c) a expression termination sequence having a nucleic acid         sequence which hybridizes under stringent conditions to a         nucleic acid sequence as shown in any one of SEQ ID NOs: 2, 7,         10, 15, 17, 23, 71, 78, 86, 96, 104, 112, or 125;     -   (d) a expression termination sequence having a nucleic acid         sequence which hybridizes to a nucleic acid sequences located         downstream of an open reading frame sequence shown in any one of         SEQ ID NOs: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112 or         125;     -   (e) a expression termination sequence having a nucleic acid         sequence which hybridizes to a nucleic acid sequences located         downstream of an open reading frame sequence being at least 80%         identical to an open reading frame sequence as shown in any one         of SEQ ID NOs: 5, 13, 20, 21, 27, 28, 71, 78, 86, 96, 104, 112         or 125;     -   (f) a expression termination sequence obtainable by 3′ genome         walking or TAIL PCR on genomic DNA from the last exon of an open         reading frame sequence as shown in any one of SEQ ID NOs: 5, 13,         20, 21, 27, 28, 71, 78, 86, 96, 104, 112 or 125; and     -   (g) a expression termination sequence obtainable by 3′ genome         walking or TAIL PCR on genomic DNA from the last exon of an open         reading frame sequence being at least 80% identical to an open         reading frame as shown in any one of SEQ ID NOs: 5, 13, 20, 21,         27, 28, 71, 78, 86, 96, 104, 112 or 125.

The term “expression termination sequence” as used herein refers to a nucleic acid which is capable of governing the termination of the process of RNA transcription of a nucleic acid operatively linked thereto, e.g. a nucleic acid of interest referred to elsewhere in this specification in detail. A termination sequence as referred to in accordance with the present invention, preferably, contains a polyadenylation signal and furthermore mediates dissociation of RNA polymerases, preferably, RNA polymerase I, II or III, more preferably, RNA polymerase II or III, and most preferably, RNA polymerase II from the transcribed DNA. Thereby the elongation of a RNA transcript, transcribed from a nucleic acid operatively linked to the termination sequence will be terminated and the RNA will be released. In order to govern termination of transcription of a nucleic acid, the expression control sequence may be located immediately adjacent to the nucleic acid whose expression is to be terminated, i.e. physically linked to the said nucleic acid at its 3″end. Alternatively, it may be located in physical proximity. In the latter case, however, the sequence must be located so as to allow functional interaction with the nucleic acid whose transcription is to be terminated. A termination sequence referred to herein, preferably, comprises between 50 and 2,000 nucleotides in length. More preferably, it comprises between 100 and 800 nucleotides and, more preferably, at least 100 nucleotides. Preferred expression termination sequences are those comprised by the polynucleotide referred to above.

Furthermore, the definitions and explanations of the terms made above apply mutatis muandis except as specified herein below.

For termination sequences, the term “operatively linked” means that the termination sequence of the present invention and a nucleic acid of interest, are linked so that the termination of transcription of the mRNA can be governed by said termination sequence, i.e. the termination sequence shall be functionally linked to said nucleic acid sequence whose transcription is to be terminated. Accordingly, the expression control sequence, the nucleic acid sequence to be expressed and the termination sequence may be physically linked to each other, e.g., by inserting the expression control sequence at the 5″end of the nucleic acid sequence to be expressed and/or inserting the termination sequence at the 3′ end of the nucleic acid sequence whose transcription is to be terminated. Alternatively, the expression control sequence and the nucleic acid to be expressed may be merely in physical proximity so that the expression control sequence is capable of governing the expression of at least one nucleic acid sequence of interest. The termination sequence and the nucleic acid whose transcription is to be terminated are, preferably, separated by not more than 50 bp, 40 bp, 20 bp, 10 bp or 5 bp.

Advantageously, the polynucleotide of the present invention comprising a expression termination sequence can be also applied for efficient expression control in plants and, in particular, plant seeds. Specifically, the expression termination sequence allows for accurate termination of transcription of the DNA into RNA after the nucleic acid sequence of interest has been transcribed. Thus, the transcription of undesired nucleic acid sequences is avoided.

The present invention also relates to a vector comprising the polynucleotide of the present invention.

The term “vector”, preferably, encompasses phage, plasmid, viral or retroviral vectors as well as artificial chromosomes, such as bacterial or yeast artificial chromosomes. Moreover, the term also relates to targeting constructs which allow for random or site-directed integration of the targeting construct into genomic DNA. Such target constructs, preferably, comprise DNA of sufficient length for either homologous or heterologous recombination as described in detail below. The vector encompassing the polynucleotides of the present invention, preferably, further comprises selectable markers for propagation and/or selection in a host. The vector may be incorporated into a host cell by various techniques well known in the art. If introduced into a host cell, the vector may reside in the cytoplasm or may be incorporated into the genome. In the latter case, it is to be understood that the vector may further comprise nucleic acid sequences which allow for homologous recombination or heterologous insertion. Vectors can be introduced into prokaryotic or eukaryotic cells via conventional transformation or transfection techniques. The terms “transformation” and “transfection”, conjugation and transduction, as used in the present context, are intended to comprise a multiplicity of prior-art processes for introducing foreign nucleic acid (for example DNA) into a host cell, including calcium phosphate, rubidium chloride or calcium chloride coprecipitation, DEAE-dextran-mediated transfection, lipofection, natural competence, carbon-based clusters, chemically mediated transfer, electroporation or particle bombardment (e.g., “gene-gun”). Suitable methods for the transformation or transfection of host cells, including plant cells, can be found in Sambrook et al. (Molecular Cloning: A Laboratory Manual, 2^(nd) ed., Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1989) and other laboratory manuals, such as Methods in Molecular Biology, 1995, Vol. 44, Agrobacterium protocols, Ed.: Gartland and Davey, Humana Press, Totowa, N.J. Alternatively, a plasmid vector may be introduced by heat shock or electroporation techniques. Should the vector be a virus, it may be packaged in vitro using an appropriate packaging cell line prior to application to host cells. Retroviral vectors may be replication competent or replication defective. In the latter case, viral propagation generally will occur only in complementing host/cells.

Preferably, the vector referred to herein is suitable as a cloning vector, i.e. replicable in microbial systems. Such vectors ensure efficient cloning in bacteria and, preferably, yeasts or fungi and make possible the stable transformation of plants. Those which must be mentioned are, in particular, various binary and co-integrated vector systems which are suitable for the T-DNA-mediated transformation. Such vector systems are, as a rule, characterized in that they contain at least the vir genes, which are required for the Agrobacterium-mediated transformation, and the sequences which delimit the T-DNA (T-DNA border). These vector systems, preferably, also comprise further cis-regulatory regions such as promoters and terminators and/or selection markers with which suitable transformed host cells or organisms can be identified. While co-integrated vector systems have vir genes and T-DNA sequences arranged on the same vector, binary systems are based on at least two vectors, one of which bears vir genes, but no T-DNA, while a second one bears T-DNA, but no vir gene. As a consequence, the last-mentioned vectors are relatively small, easy to manipulate and can be replicated both in E. coli and in Agrobacterium. These binary vectors include vectors from the pBIB-HYG, pPZP, pBecks, pGreen series. Preferably used in accordance with the invention are Bin19, pBUI101, pBinAR, pGPTV, pSUN and pCAMBIA. An overview of binary vectors and their use can be found in Hellens et al, Trends in Plant Science (2000) 5, 446-451. Furthermore, by using appropriate cloning vectors, the polynucleotide of the invention can be introduced into host cells or organisms such as plants or animals and, thus, be used in the transformation of plants, such as those which are published, and cited, in: Plant Molecular Biology and Biotechnology (CRC Press, Boca Raton, Fla.), chapter 6/7, pp. 71-119 (1993); F. F. White, Vectors for Gene Transfer in Higher Plants; in: Transgenic Plants, vol. 1, Engineering and Utilization, Ed.: Kung and R. Wu, Academic Press, 1993, 15-38; B. Jenes et al., Techniques for Gene Transfer, in: Transgenic Plants, vol. 1, Engineering and Utilization, Ed.: Kung and R. Wu, Academic Press (1993), 128-143; Potrykus, Annu. Rev. Plant Physiol. Plant Molec. Biol. 42 (1991), 205-225.

More preferably, the vector of the present invention is an expression vector. In such an expression vector, the polynucleotide comprises an expression cassette as specified above allowing for expression in eukaryotic cells or isolated fractions thereof. An expression vector may, in addition to the polynucleotide of the invention, also comprise further regulatory elements including transcriptional as well as translational enhancers. Preferably, the expression vector is also a gene transfer or targeting vector. Expression vectors derived from viruses such as retroviruses, vaccinia virus, adeno-associated virus, herpes viruses, or bovine papilloma virus, may be used for delivery of the polynucleotides or vector of the invention into targeted cell population. Methods which are well known to those skilled in the art can be used to construct recombinant viral vectors; see, for example, the techniques described in Sambrook, Molecular Cloning A Laboratory Manual, Cold Spring Harbor Laboratory (1989) N.Y. and Ausubel, Current Protocols in Molecular Biology, Green Publishing Associates and Wiley Interscience, N.Y. (1994).

Suitable expression vector backbones are, preferably, derived from expression vectors known in the art such as Okayama-Berg cDNA expression vector pcDV1 (Pharmacia), pCDM8, pRc/CMV, pcDNA1, pcDNA3 (Invitrogene) or pSPORT1 (GIBCO BRL). Further examples of typical fusion expression vectors are pGEX (Pharmacia Biotech Inc; Smith, D. B., and Johnson, K. S. (1988) Gene 67:31-40), pMAL (New England Biolabs, Beverly, Mass.) and pRIT5 (Pharmacia, Piscataway, N.J.), where glutathione S-transferase (GST), maltose E-binding protein and protein A, respectively, are fused with the nucleic acid of interest encoding a protein to be expressed. The target gene expression of the pTrc vector is based on the transcription from a hybrid trp-lac fusion promoter by host RNA polymerase. The target gene expression from the pET 11d vector is based on the transcription of a T7-gn10-lac fusion promoter, which is mediated by a coexpressed viral RNA polymerase (T7 gn1). This viral polymerase is provided by the host strains BL21 (DE3) or HMS174 (DE3) from a resident λ-prophage which harbors a T7 gn1 gene under the transcriptional control of the lacUV 5 promoter. Examples of vectors for expression in the yeast S. cerevisiae comprise pYepSec1 (Baldari et al. (1987) Embo J. 6:229-234), pMFa (Kurjan and Herskowitz (1982) Cell 30:933-943), pJRY88 (Schultz et al. (1987) Gene 54:113-123) and pYES2 (Invitrogen Corporation, San Diego, Calif.). Vectors and processes for the construction of vectors which are suitable for use in other fungi, such as the filamentous fungi, comprise those which are described in detail in: van den Hondel, C. A. M. J. J., & Punt, P. J. (1991) “Gene transfer systems and vector development for filamentous fungi, in: Applied Molecular Genetics of fungi, J. F. Peberdy et al., Ed., pp. 1-28, Cambridge University Press: Cambridge, or in: More Gene Manipulations in Fungi (J. W. Bennett & L. L. Lasure, Ed., pp. 396-428: Academic Press: San Diego). Further suitable yeast vectors are, for example, pAG-1, YEp6, YEp13 or pEMBLYe23. As an alternative, the polynucleotides of the present invention can be also expressed in insect cells using baculovirus expression vectors. Baculovirus vectors which are available for the expression of proteins in cultured insect cells (for example Sf9 cells) comprise the pAc series (Smith et al. (1983) Mol. Cell. Biol. 3:2156-2165) and the pVL series (Lucklow and Summers (1989) Virology 170:31-39).

The polynucleotides of the present invention can be used for expression of a nucleic acid of interest in single-cell plant cells (such as algae), see Falciatore et al., 1999, Marine Biotechnology 1 (3):239-251 and the references cited therein, and plant cells from higher plants (for example Spermatophytes, such as arable crops) by using plant expression vectors. Examples of plant expression vectors comprise those which are described in detail in: Becker, D., Kemper, E., Schell, J., and Masterson, R. (1992) “New plant binary vectors with selectable markers located proximal to the left border”, Plant Mol. Biol. 20:1195-1197; and Bevan, M. W. (1984) “Binary Agrobacterium vectors for plant transformation”, Nucl. Acids Res. 12:8711-8721; Vectors for Gene Transfer in Higher Plants; in: Transgenic Plants, Vol. 1, Engineering and Utilization, Ed.: Kung and R. Wu, Academic Press, 1993, p. 15-38. A plant expression cassette, preferably, comprises regulatory sequences which are capable of controlling the gene expression in plant cells and which are functionally linked so that each sequence can fulfill its function, such as transcriptional termination, for example polyadenylation signals. Preferred polyadenylation signals are those which are derived from Agrobacterium tumefaciens T-DNA, such as the gene 3 of the Ti plasmid pTiACH5, which is known as octopine synthase (Gielen et al., EMBO J. 3 (1984) 835 et seq.) or functional equivalents of these, but all other terminators which are functionally active in plants are also suitable. Since plant gene expression is very often not limited to transcriptional levels, a plant expression cassette preferably comprises other functionally linked sequences such as translation enhancers, for example the overdrive sequence, which comprises the 5′-untranslated tobacco mosaic virus leader sequence, which increases the protein/RNA ratio (Gallie et al., 1987, Nucl. Acids Research 15:8693-8711). Other preferred sequences for the use in functional linkage in plant gene expression cassettes are targeting sequences which are required for targeting the gene product into its relevant cell compartment (for a review, see Kermode, Crit. Rev. Plant Sci. 15, 4 (1996) 285-423 and references cited therein), for example into the vacuole, the nucleus, all types of plastids, such as amyloplasts, chloroplasts, chromoplasts, the extracellular space, the mitochondria, the endoplasmic reticulum, oil bodies, peroxisomes and other compartments of plant cells.

The abovementioned vectors are only a small overview of vectors to be used in accordance with the present invention. Further vectors are known to the skilled worker and are described, for example, in: Cloning Vectors (Ed., Pouwels, P. H., et al., Elsevier, Amsterdam-New York-Oxford, 1985, ISBN 0 444 904018). For further suitable expression systems for prokaryotic and eukaryotic cells see the chapters 16 and 17 of Sambrook, J., Fritsch, E. F., and Maniatis, T., Molecular Cloning: A Laboratory Manual, 2^(nd) edition, Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1989.

The present invention also contemplates a host cell comprising the polynucleotide or the vector of the present invention.

Host cells are primary cells or cell lines derived from multicellular organisms such as plants or animals. Furthermore, host cells encompass prokaryotic or eukaryotic single cell organisms (also referred to as micro-organisms). Primary cells or cell lines to be used as host cells in accordance with the present invention may be derived from the multicellular organisms referred to below. Host cells which can be exploited are furthermore mentioned in: Goeddel, Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, Calif. (1990). Specific expression strains which can be used, for example those with a lower protease activity, are described in: Gottesman, S., Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, Calif. (1990) 119-128. These include plant cells and certain tissues, organs and parts of plants in all their phenotypic forms such as anthers, fibers, root hairs, stalks, embryos, calli, cotelydons, petioles, harvested material, plant tissue, reproductive tissue and cell cultures which are derived from the actual transgenic plant and/or can be used for bringing about the transgenic plant. Preferably, the host cells may be obtained from plants. More preferably, oil crops are envisaged which comprise large amounts of lipid compounds, such as oilseed rape, evening primrose, hemp, thistle, peanut, canola, linseed, soybean, safflower, sunflower, borage, or plants such as maize, wheat, rye, oats, triticale, rice, barley, cotton, cassava, pepper, Tagetes, Solanaceae plants such as potato, tobacco, eggplant and tomato, Vicia species, pea, alfalfa, bushy plants (coffee, cacao, tea), Salix species, trees (oil palm, coconut) and perennial grasses and fodder crops. Especially preferred plants according to the invention are oil crops such as soybean, peanut, oilseed rape, canola, linseed, hemp, evening primrose, sunflower, safflower, trees (oil palm, coconut). Suitable methods for obtaining host cells from the multicellular organisms referred to below as well as conditions for culturing these cells are well known in the art.

The micro-organisms are, preferably, bacteria or fungi including yeasts. Preferred fungi to be used in accordance with the present invention are selected from the group of the families Chaetomiaceae, Choanephoraceae, Cryptococcaceae, Cunninghamellaceae, Demetiaceae, Moniliaceae, Mortierellaceae, Mucoraceae, Pythiaceae, Sacharomycetaceae, Saprolegniaceae, Schizosacharomycetaceae, Sodariaceae or Tuberculariaceae. Further preferred micro-organisms are selected from the group: Choanephoraceae such as the genera Blakeslee, Choanephora, for example the genera and species Blakeslea trispora, Choanephora cucurbitarum, Choanephora infundibulifera var. cucurbitarum, Mortierellaceae, such as the genus Mortierella, for example the genera and species Mortierella isabeffina, Mortierella polycephala, Mortierella ramanniana, Mortierella vinacea, Mortierella zonata, Pythiaceae such as the genera Phytium, hytophthora for example the genera and species Pythium debaryanum, Pythium intermedium, Pythium irregulare, Pythium megalacanthum, Pythium paroecandrum, Pythium sylvaticum, Pythium ultimum, Phytophthora cactorum, Phytophthora cinnamomi, Phytophthora citricola, Phytophthora citrophthora, Phytophthora cryptogea, Phytophthora drechsleri, Phytophthora erythroseptica, Phytophthora lateralis, Phytophthora megasperma, Phytophthora nicotianae, Phytophthora nicotianae var. parasitica, Phytophthora palmivora, Phytophthora parasitica, Phytophthora syringae, Saccharomycetaceae such as the genera Hansenula, Pichia, Saccharomyces, Saccharomycodes, Yarrowia for example the genera and species Hansenula anomala, Hansenula californica, Hansenula canadensis, Hansenula capsulata, Hansenula ciferrii, Hansenula glucozyma, Hansenula henricii, Hansenula holstii, Hansenula minuta, Hansenula nonfermentans, Hansenula philodendri, Hansenula polymorpha, Hansenula saturnus, Hansenula subpelliculosa, Hansenula wickerhamii, Hansenula wingei, Pichia alcoholophila, Pichia angusta, Pichia anomala, Pichia bispora, Pichia burtonii, Pichia canadensis, Pichia capsulata, Pichia carsonii, Pichia cellobiosa, Pichia ciferrii, Pichia farinosa, Pichia fermentans, Pichia finlandica, Pichia glucozyma, Pichia guilliermondii, Pichia haplophila, Pichia henricii, Pichia holstii, Pichia jadinii, Pichia lindnerii, Pichia membranaefaciens, Pichia methanolica, Pichia minuta var. minuta, Pichia minuta var. nonfermentans, Pichia norvegensis, Pichia ohmeri, Pichia pastoris, Pichia philodendri, Pichia pini, Pichia polymorpha, Pichia quercuum, Pichia rhodanensis, Pichia sargentensis, Pichia stipitis, Pichia strasburgensis, Pichia subpelliculosa, Pichia toletana, Pichia trehalophila, Pichia vini, Pichia xylosa, Saccharomyces aceta, Saccharomyces bailii, Saccharomyces bayanus, Saccharomyces bisporus, Saccharomyces capensis, Saccharomyces carlsbergensis, Saccharomyces cerevisiae, Saccharomyces cerevisiae var. ellipsoideus, Saccharomyces chevalieri, Saccharomyces delbrueckii, Saccharomyces diastaticus, Saccharomyces drosophilarum, Saccharomyces elegans, Saccharomyces ellipsoideus, Saccharomyces fermentati, Saccharomyces florentinus, Saccharomyces fragilis, Saccharomyces heterogenicus, Saccharomyces hienipiensis, Saccharomyces inusitatus, Saccharomyces italicus, Saccharomyces kluyveri, Saccharomyces krusei, Saccharomyces lactis, Saccharomyces marxianus, Saccharomyces microellipsoides, Saccharomyces montanus, Saccharomyces norbensis, Saccharomyces oleaceus, Saccharomyces paradoxus, Saccharomyces pastorianus, Saccharomyces pretoriensis, Saccharomyces rosea, Saccharomyces rouxii, Saccharomyces uvarum, Saccharomycodes ludwigii, Yarrowia lipolytica, Schizosacharomycetaceae such as the genera Schizosaccharomyces e.g. the species Schizosaccharomyces japonicus var. japonicus, Schizosaccharomyces japonicus var. versatilis, Schizosaccharomyces malidevorans, Schizosaccharomyces octosporus, Schizosaccharomyces pombe var. malidevorans, Schizosaccharomyces pombe var. pombe, Thraustochytriaceae such as the genera Althornia, Aplanochytrium, Japonochytrium, Schizochytrium, Thraustochytrium e.g. the species Schizochytrium aggregatum, Schizochytrium limacinum, Schizochytrium mangrovei, Schizochytrium minutum, Schizochytrium octosporum, Thraustochytrium aggregatum, Thraustochytrium amoeboideum, Thraustochytrium antacticum, Thraustochytrium arudimentale, Thraustochytrium aureum, Thraustochytrium benthicola, Thraustochytrium globosum, Thraustochytrium indicum, Thraustochytrium kerguelense, Thraustochytrium kinnei, Thraustochytrium motivum, Thraustochytrium multirudimentale, Thraustochytrium pachydermum, Thraustochytrium proliferum, Thraustochytrium roseum, Thraustochytrium rossii, Thraustochytrium striatum or Thraustochytrium visurgense. Further preferred microorganisms are bacteria selected from the group of the families Bacillaceae, Enterobacteriacae or Rhizobiaceae. Examples of such micro-organisms may be selected from the group: Bacillaceae such as the genera Bacillus for example the genera and species Bacillus acidocaldarius, Bacillus acidoterrestris, Bacillus alcalophilus, Bacillus amyloliquefaciens, Bacillus amylolyticus, Bacillus brevis, Bacillus cereus, Bacillus circulans, Bacillus coagulans, Bacillus sphaericus subsp. fusiformis, Bacillus galactophilus, Bacillus globisporus, Bacillus globisporus subsp. marinus, Bacillus halophilus, Bacillus lentimorbus, Bacillus lentus, Bacillus licheniformis, Bacillus megaterium, Bacillus polymyxa, Bacillus psychrosaccharolyticus, Bacillus pumilus, Bacillus sphaericus, Bacillus subtilis subsp. spizizenii, Bacillus subtilis subsp. subtilis or Bacillus thuringiensis; Enterobacteriacae such as the genera Citrobacter, Edwardsiella, Enterobacter, Erwinia, Escherichia, Klebsiella, Salmonella or Serratia for example the genera and species Citrobacter amalonaticus, Citrobacter diversus, Citrobacter freundii, Citrobacter genomospecies, Citrobacter gillenii, Citrobacter intermedium, Citrobacter koseri, Citrobacter murliniae, Citrobacter sp., Edwardsiella hoshinae, Edwardsiella ictaluri, Edwardsiella tarda, Erwinia alni, Erwinia amylovora, Erwinia ananatis, Erwinia aphidicola, Erwinia billingiae, Erwinia cacticida, Erwinia cancerogena, Erwinia carnegieana, Erwinia carotovora subsp. atroseptica, Erwinia carotovora subsp. betavasculorum, Erwinia carotovora subsp. odorifera, Erwinia carotovora subsp. wasabiae, Erwinia chrysanthemi, Erwinia cypripedii, Erwinia dissolvens, Erwinia herbicola, Erwinia mallotivora, Erwinia milletiae, Erwinia nigrifluens, Erwinia nimipressuralis, Erwinia persicina, Erwinia psidii, Erwinia pyrifoliae, Erwinia quercina, Erwinia rhapontici, Erwinia rubrifaciens, Erwinia salicis, Erwinia stewartii, Erwinia tracheiphila, Erwinia uredovora, Escherichia adecarboxylata, Escherichia anindolica, Escherichia aurescens, Escherichia blattae, Escherichia coli, Escherichia coli var. communion, Escherichia coli-mutabile, Escherichia fergusonii, Escherichia hermannii, Escherichia sp., Escherichia vulneris, Klebsiella aerogenes, Klebsiella edwardsii subsp. atlantae, Klebsiella ornithinolytica, Klebsiella oxytoca, Klebsiella planticola, Klebsiella pneumoniae, Klebsiella pneumoniae subsp. pneumoniae, Klebsiella sp., Klebsiella terrigena, Klebsiella trevisanii, Salmonella abony, Salmonella arizonae, Salmonella bongori, Salmonella choleraesuis subsp. arizonae, Salmonella choleraesuis subsp. bongori, Salmonella choleraesuis subsp. cholereasuis, Salmonella choleraesuis subsp. diarizonae, Salmonella choleraesuis subsp. houtenae, Salmonella choleraesuis subsp. indica, Salmonella choleraesuis subsp. salamae, Salmonella daressalaam, Salmonella enterica subsp. houtenae, Salmonella enterica subsp. salamae, Salmonella enteritidis, Salmonella gallinarum, Salmonella heidelberg, Salmonella panama, Salmonella senftenberg, Salmonella typhimurium, Serratia entomophila, Serratia ficaria, Serratia fonticola, Serratia grimesii, Serratia liquefaciens, Serratia marcescens, Serratia marcescens subsp. marcescens, Serratia marinorubra, Serratia odorifera, Serratia plymouthensis, Serratia plymuthica, Serratia proteamaculans, Serratia proteamaculans subsp. quinovora, Serratia quinivorans or Serratia rubidaea; Rhizo-biaceae such as the genera Agrobacterium, Carbophilus, Chelatobacter, Ensifer, Rhizobium, Sinorhizobium for example the genera and species Agrobacterium atlanticum, Agrobacterium ferrugineum, Agrobacterium gelatinovorum, Agrobacterium larrymoorei, Agrobacterium meteori, Agrobacterium radiobacter, Agrobacterium rhizogenes, Agrobacterium rubi, Agrobacterium stellulatum, Agrobacterium tumefaciens, Agrobacterium vitis, Carbophilus carboxidus, Chelatobacter heintzii, Ensifer adhaerens, Ensifer arboris, Ensifer fredii, Ensifer kostiensis, Ensifer kummerowiae, Ensifer medicae, Ensifer meliloti, Ensifer saheli, Ensifer terangae, Ensifer xinjiangensis, Rhizobium ciceri Rhizobium etli, Rhizobium fredii, Rhizobium galegae, Rhizobium gallicum, Rhizobium giardinii, Rhizobium hainanense, Rhizobium huakuii, Rhizobium huautlense, Rhizobium indigoferae, Rhizobium japonicum, Rhizobium leguminosarum, Rhizobium loessense, Rhizobium loti, Rhizobium lupini, Rhizobium mediterraneum, Rhizobium meliloti, Rhizobium mongolense, Rhizobium phaseoli, Rhizobium radiobacter, Rhizobium rhizogenes, Rhizobium rubi, Rhizobium sullae, Rhizobium tianshanense, Rhizobium trifolii, Rhizobium tropici, Rhizobium undicola, Rhizobium vitis, Sinorhizobium adhaerens, Sinorhizobium arboris, Sinorhizobium fredii, Sinorhizobium kostiense, Sinorhizobium kummerowiae, Sinorhizobium medicae, Sinorhizobium meliloti, Sinorhizobium morelense, Sinorhizobium saheli or Sinorhizobium xinjiangense.

How to culture the aforementioned micro-organisms is well known to the person skilled in the art.

The present invention also relates to a non-human transgenic organism, preferably a plant or seed thereof, comprising the polynucleotide or the vector of the present invention.

The term “non-human transgenic organism”, preferably, relates to a plant, a plant seed, a non-human animal or a multicellular micro-organism. The polynucleotide or vector may be present in the cytoplasm of the organism or may be incorporated into the genome either heterologous or by homologous recombination. Host cells, in particular those obtained from plants or animals, may be introduced into a developing embryo in order to obtain mosaic or chimeric organisms, i.e. non-human transgenic organisms comprising the host cells of the present invention. Suitable transgenic organisms are, preferably, all organisms which are suitable for the expression of recombinant genes. Preferred plants to be used for making non-human transgenic organisms according to the present invention are all dicotyledonous or monocotyledonous plants, algae or mosses. Advantageous plants are selected from the group of the plant families Adelotheciaceae, Anacardiaceae, Asteraceae, Apiaceae, Betulaceae, Boraginaceae, Brassicaceae, Bromeliaceae, Caricaceae, Cannabaceae, Convolvulaceae, Chenopodiaceae, Crypthecodiniaceae, Cucurbitaceae, Ditrichaceae, Elaeagnaceae, Ericaceae, Euphorbiaceae, Fabaceae, Geraniaceae, Gramineae, Juglandaceae, Lauraceae, Leguminosae, Linaceae, Prasinophyceae or vegetable plants or ornamentals such as Tagetes. Examples which may be mentioned are the following plants selected from the group consisting of: Adelotheciaceae such as the genera Physcomitrella, such as the genus and species Physcomitrella patens, Anacardiaceae such as the genera Pistacia, Mangifera, Anacardium, for example the genus and species Pistacia vera [pistachio], Mangifer indica [mango] or Anacardium occidentale [cashew], Asteraceae, such as the genera Calendula, Carthamus, Centaurea, Cichorium, Cynara, Helianthus, Lactuca, Locusta, Tagetes, Valeriana, for example the genus and species Calendula offinalis [common marigold], Carthamus tinctorius [safflower], Centaurea cyanus [cornflower], Cichorium intybus [chicory], Cynara scolymus [artichoke], Helianthus annus [sunflower], Lactuca sativa, Lactuca crispa, Lactuca esculenta, Lactuca scariola L. ssp. sativa, Lactuca scariola L. var. integrate, Lactuca scariola L. var. integrifolia, Lactuca sativa subsp. romana, Locusta communis, Valeriana locusta [salad vegetables], Tagetes lucida, Tagetes erecta or Tagetes tenuifolia [african or french marigold], Apiaceae, such as the genus Daucus, for example the genus and species Daucus carota [carrot], Betulaceae, such as the genus Corylus, for example the genera and species Corylus avellana or Corylus colurna [hazelnut], Boraginaceae, such as the genus Borago, for example the genus and species Borago officinalis [borage], Brassicaceae, such as the genera Brassica, Melanosinapis, Sinapis, Arabadopsis, for example the genera and species Brassica napus, Brassica rapa ssp. [oilseed rape], Sinapis arvensis Brassica juncea, Brassica juncea var. juncea, Brassica juncea var. crispifolia, Brassica juncea var. foliosa, Brassica nigra, Brassica sinapioides, Melanosinapis communis [mustard], Brassica oleracea [fodder beet] or Arabidopsis thaliana, Bromeliaceae, such as the genera Anana, Bromelia (pineapple), for example the genera and species Anana comosus, Ananas ananas or Bromelia comosa [pineapple], Caricaceae, such as the genus Carica, such as the genus and species Carica papaya [pawpaw], Cannabaceae, such as the genus Cannabis, such as the genus and species Cannabis sativa [hemp], Convolvulaceae, such as the genera Ipomea, Convolvulus, for example the genera and species Ipomoea batatus, Ipomoea pandurata, Convolvulus batatas, Convolvulus tiliaceus, Ipomoea fastigiata, Ipomoea tiliacea, Ipomoea triloba or Convolvulus panduratus [sweet potato, batate], Chenopodiaceae, such as the genus Beta, such as the genera and species Beta vulgaris, Beta vulgaris var. altissima, Beta vulgaris var. Vulgaris, Beta maritima, Beta vulgaris var. perennis, Beta vulgaris var. conditiva or Beta vulgaris var. esculenta [sugarbeet], Crypthecodiniaceae, such as the genus Crypthecodinium, for example the genus and species Cryptecodinium cohnii, Cucurbitaceae, such as the genus Cucurbita, for example the genera and species Cucurbita maxima, Cucurbita mixta, Cucurbita pepo or Cucurbita moschata [pumpkin/squash], Cymbellaceae such as the genera Amphora, Cymbella, Okedenia, Phaeodactylum, Reimeria, for example the genus and species Phaeodactylum tricornutum, Ditrichaceae such as the genera Ditrichaceae, Astomiopsis, Ceratodon, Chrysoblastella, Ditrichum, Distichium, Eccremidium, Lophidion, Philibertiella, Pleuridium, Saelania, Trichodon, Skottsbergia, for example the genera and species Ceratodon antarcticus, Ceratodon columbiae, Ceratodon heterophyllus, Ceratodon purpureus, Ceratodon purpureus, Ceratodon purpureus ssp. convolutus, Ceratodon, purpureus spp. stenocarpus, Ceratodon purpureus var. rotundifolius, Ceratodon ratodon, Ceratodon stenocarpus, Chrysoblastella chilensis, Ditrichum ambiguum, Ditrichum brevisetum, Ditrichum crispatissimum, Ditrichum difficile, Ditrichum falcifolium, Ditrichum flexicaule, Ditrichum giganteum, Ditrichum heteromallum, Ditrichum lineare, Ditrichum lineare, Ditrichum montanum, Ditrichum montanum, Ditrichum pallidum, Ditrichum punctulatum, Ditrichum pusillum, Ditrichum pusillum var. tortile, Ditrichum rhynchostegium, Ditrichum schimperi, Ditrichum tortile, Distichium capillaceum, Distichium hagenii, Distichium inclinatum, Distichium macounii, Eccremidium floridanum, Eccremidium whiteleggei, Lophidion strictus, Pleuridium acuminatum, Pleuridium alternifolium, Pleuridium holdridgei, Pleuridium mexicanum, Pleuridium ravenelii, Pleuridium subulatum, Saelania glaucescens, Trichodon borealis, Trichodon cylindricus or Trichodon cylindricus var. oblongus, Elaeagnaceae such as the genus Elaeagnus, for example the genus and species Olea europaea [olive], Ericaceae such as the genus Kalmia, for example the genera and species Kalmia latifolia, Kalmia angustifolia, Kalmia microphylla, Kalmia polifolia, Kalmia occidentalis, Cistus chamaerhodendros or Kalmia lucida [mountain laurel], Euphorbiaceae such as the genera Manihot, Janipha, Jatropha, Ricinus, for example the genera and species Manihot utilissima, Janipha manihot, Jatropha manihot, Manihot aipil, Manihot dulcis, Manihot manihot, Manihot melanobasis, Manihot esculenta [manihot] or Ricinus communis [castor-oil plant], Fabaceae such as the genera Pisum, Albizia, Cathormion, Feuillea, Inga, Pithecolobium, Acacia, Mimosa, Medicajo, Glycine, Dolichos, Phaseolus, Soja, for example the genera and species Pisum sativum, Pisum arvense, Pisum humile [pea], Albizia berteriana, Albizia julibrissin, Albizia lebbeck, Acacia berteriana, Acacia littoralis, Albizia berteriana, Albizzia berteriana, Cathormion berteriana, Feuillea berteriana, Inga fragrans, Pithecellobium berterianum, Pithecellobium fragrans, Pithecolobium berterianum, Pseudalbizzia berteriana, Acacia julibrissin, Acacia nemu, Albizia nemu, Feuilleea julibrissin, Mimosa julibrissin, Mimosa speciosa, Sericanrda julibrissin, Acacia lebbeck, Acacia macrophylla, Albizia lebbek, Feuilleea lebbeck, Mimosa lebbeck, Mimosa speciosa [silk tree], Medicago sativa, Medicago falcata, Medicago varia [alfalfa], Glycine max Dolichos soja, Glycine gracilis, Glycine hispida, Phaseolus max, Soja hispida or Soja max [soybean], Funariaceae such as the genera Aphanorrhegma, Entosthodon, Funaria, Physcomitrella, Physcomitrium, for example the genera and species Aphanorrhegma serratum, Entosthodon attenuatus, Entosthodon bolanderi, Entosthodon bonplandii, Entosthodon californicus, Entosthodon drummondii, Entosthodon jamesonii, Entosthodon leibergii, Entosthodon neoscoticus, Entosthodon rubrisetus, Entosthodon spathulifolius, Entosthodon tucsoni, Funaria americana, Funaria bolanderi, Funaria calcarea, Funaria californica, Funaria calvescens, Funaria convoluta, Funaria flavicans, Funaria groutiana, Funaria hygrometrica, Funaria hygrometrica var. arctica, Funaria hygrometrica var. calvescens, Funaria hygrometrica var. convoluta, Funaria hygrometrica var. muralis, Funaria hygrometrica var. utahensis, Funaria microstoma, Funaria microstoma var. obtusifolia, Funaria muhlenbergii, Funaria orcuttii, Funaria plano-convexa, Funaria polaris, Funaria ravenelii, Funaria rubriseta, Funaria serrata, Funaria sonorae, Funaria sublimbatus, Funaria tucsoni, Physcomitrella californica, Physcomitrella patens, Physcomitrella readeri, Physcomitrium australe, Physcomitrium californicum, Physcomitrium collenchymatum, Physcomitrium coloradense, Physcomitrium cupuliferum, Physcomitrium drummondii, Physcomitrium eurystomum, Physcomitrium flexifolium, Physcomitrium hookeri, Physcomitrium hookeri var. serratum, Physcomitrium immersum, Physcomitrium kellermanii, Physcomitrium megalocarpum, Physcomitrium pyriforme, Physcomitrium pyriforme var. serratum, Physcomitrium rufipes, Physcomitrium sandbergii, Physcomitrium subsphaericum, Physcomitrium washingtoniense, Geraniaceae, such as the genera Pelargonium, Cocos, Oleum, for example the genera and species Cocos nucifera, Pelargonium grossularioides or Oleum cocois [coconut], Gramineae, such as the genus Saccharum, for example the genus and species Saccharum officinarum, Juglandaceae, such as the genera Juglans, Wallia, for example the genera and species Juglans regia, Juglans ailanthifolia, Juglans sieboldiana, Juglans cinerea, Wallia cinerea, Juglans bixbyi, Juglans californica, Juglans hindsii, Juglans intermedia, Juglans jamaicensis, Juglans major, Juglans microcarpa, Juglans nigra or Wallia nigra [walnut], Lauraceae, such as the genera Persea, Laurus, for example the genera and species Laurus nobilis [bay], Persea americana, Persea gratissima or Persea persea [avocado], Leguminosae, such as the genus Arachis, for example the genus and species Arachis hypogaea [peanut], Linaceae, such as the genera Linum, Adenolinum, for example the genera and species Linum usitatissimum, Linum humile, Linum austriacum, Linum bienne, Linum angustifolium, Linum catharticum, Linum flavum, Linum grandiflorum, Adenolinum grandiflorum, Linum lewisii, Linum narbonense, Linum perenne, Linum perenne var. lewisii, Linum pratense or Linum trigynum [linseed], Lythrarieae, such as the genus Punica, for example the genus and species Punica granatum [pomegranate], Malvaceae, such as the genus Gossypium, for example the genera and species Gossypium hirsutum, Gossypium arboreum, Gossypium barbadense, Gossypium herbaceum or Gossypium thurberi [cotton], Marchantiaceae, such as the genus Marchantia, for example the genera and species Marchantia berteroana, Marchantia foliacea, Marchantia macropora, Musaceae, such as the genus Musa, for example the genera and species Musa nana, Musa acuminata, Musa paradisiaca, Musa spp. [banana], Onagraceae, such as the genera Camissonia, Oenothera, for example the genera and species Oenothera biennis or Camissonia brevipes [evening primrose], Palmae, such as the genus Elacis, for example the genus and species Elaeis guineensis [oil palm], Papaveraceae, such as the genus Papaver, for example the genera and species Papaver orientale, Papaver rhoeas, Papaver dubium [poppy], Pedaliaceae, such as the genus Sesamum, for example the genus and species Sesamum indicum [sesame], Piperaceae, such as the genera Piper, Artanthe, Peperomia, Steffensia, for example the genera and species Piper aduncum, Piper amalago, Piper angustifolium, Piper auriturn, Piper betel, Piper cubeba, Piper longum, Piper nigrum, Piper retrofractum, Artanthe adunca, Artanthe elongata, Peperomia elongata, Piper elongatum, Steffensia elongata [cayenne pepper], Poaceae, such as the genera Hordeum, Secale, Avena, Sorghum, Andropogon, Holcus, Panicum, Oryza, Zea (maize), Triticum, for example the genera and species Hordeum vulgare, Hordeum jubatum, Hordeum murinum, Hordeum secalinum, Hordeum distichon, Hordeum aegiceras, Hordeum hexastichon, Hordeum hexastichum, Hordeum irregulare, Hordeum sativum, Hordeum secalinum [barley], Secale cereale [rye], Avena sativa, Avena fatua, Avena byzantina, Avena fatua var. sativa, Avena hybrida [oats], Sorghum bicolor, Sorghum halepense, Sorghum saccharatum, Sorghum vulgare, Andropogon drummondii, Holcus bicolor, Holcus sorghum, Sorghum aethiopicum, Sorghum arundinaceum, Sorghum caffrorum, Sorghum cernuum, Sorghum dochna, Sorghum drummondii, Sorghum durra, Sorghum guineense, Sorghum lanceolatum, Sorghum nervosum, Sorghum saccharatum, Sorghum subglabrescens, Sorghum verticilliflorum, Sorghum vulgare, Holcus halepensis, Sorghum miliaceum, Panicum militaceum [millet], Oryza sativa, Oryza latifolia [rice], Zea mays [maize], Triticum aestivum, Triticum durum, Triticum turgidum, Triticum hybernum, Triticum macha, Triticum sativum or Triticum vulgare [wheat], Porphyridiaceae, such as the genera Chroothece, Flintiella, Petrovanella, Porphyridium, Rhodella, Rhodosorus, Vanhoeffenia, for example the genus and species Porphyridium cruentum, Proteaceae, such as the genus Macadamia, for example the genus and species Macadamia intergrifolia [macadamia], Prasinophyceae such as the genera Nephroselmis, Prasinococcus, Scherffelia, Tetraselmis, Mantoniella, Ostreococcus, for example the genera and species Nephroselmis olivacea, Prasinococcus capsulatus, Scherffelia dubia, Tetraselmis chui, Tetraselmis suecica, Mantoniella squamata, Ostreococcus tauri, Rubiaceae such as the genus Cofea, for example the genera and species Cofea spp., Coffea arabica, Coffea canephora or Coffea liberica [coffee], Scrophulariaceae such as the genus Verbascum, for example the genera and species Verbascum blattaria, Verbascum chaixii, Verbascum densiflorum, Verbascum lagurus, Verbascum longifolium, Verbascum lychnitis, Verbascum nigrum, Verbascum olympicum, Verbascum phlomoides, Verbascum phoenicum, Verbascum pulverulentum or Verbascum thapsus [mullein], Solanaceae such as the genera Capsicum, Nicotiana, Solanum, Lycopersicon, for example the genera and species Capsicum annuum, Capsicum annuum var. glabriusculum, Capsicum frutescens [pepper], Capsicum annuum [paprika], Nicotiana tabacum, Nicotiana alata, Nicotiana attenuata, Nicotiana glauca, Nicotiana langsdorffii, Nicotiana obtusifolia, Nicotiana quadrivalvis, Nicotiana repanda, Nicotiana rustica, Nicotiana sylvestris [tobacco], Solanum tuberosum [potato], Solanum melongena [eggplant], Lycopersicon esculentum, Lycopersicon lycopersicum, Lycopersicon pyriforme, Solanum integrifolium or Solanum lycopersicum [tomato], Sterculiaceae, such as the genus Theobroma, for example the genus and species Theobroma cacao [cacao] or Theaceae, such as the genus Camellia, for example the genus and species Camellia sinensis [tea]. In particular preferred plants to be used as transgenic plants in accordance with the present invention are oil fruit crops which comprise large amounts of lipid compounds, such as peanut, oilseed rape, canola, sunflower, safflower, poppy, mustard, hemp, castor-oil plant, olive, sesame, Calendula, Punica, evening primrose, mullein, thistle, wild roses, hazelnut, almond, macadamia, avocado, bay, pumpkin/squash, linseed, soybean, pistachios, borage, trees (oil palm, coconut, walnut) or crops such as maize, wheat, rye, oats, triticale, rice, barley, cotton, cassava, pepper, Tagetes, Solanaceae plants such as potato, tobacco, eggplant and tomato, Vicia species, pea, alfalfa or bushy plants (coffee, cacao, tea), Salix species, and perennial grasses and fodder crops. Preferred plants according to the invention are oil crop plants such as peanut, oilseed rape, canola, sunflower, safflower, poppy, mustard, hemp, castor-oil plant, olive, Calendula, Punica, evening primrose, pumpkin/squash, linseed, soybean, borage, trees (oil palm, coconut). Especially preferred are plants which are high in C18:2- and/or C18:3-fatty acids, such as sunflower, safflower, tobacco, mullein, sesame, cotton, pumpkin/squash, poppy, evening primrose, walnut, linseed, hemp, thistle or safflower. Very especially preferred plants are plants such as safflower, sunflower, poppy, evening primrose, walnut, linseed, or hemp.

Preferred mosses are Physcomitrella or Ceratodon. Preferred algae are Isochrysis, Mantoniella, Ostreococcus or Crypthecodinium, and algae/diatoms such as Phaeodactylum or Thraustochytrium. More preferably, said algae or mosses are selected from the group consisting of: Shewanella, Physcomitrella, Thraustochytrium, Fusarium, Phytophthora, Ceratodon, lsochrysis, Aleurita, Muscarioides, Mortierella, Phaeodactylum, Cryphthecodinium, specifically from the genera and species Thallasiosira pseudonona, Euglena gracilis, Physcomitrella patens, Phytophtora infestans, Fusarium graminaeum, Cryptocodinium cohnii, Ceratodon purpureus, lsochrysis galbana, Aleurita farinosa, Thraustochytrium sp., Muscarioides viallii, Mortierella alpina, Phaeodactylum tricornutum or Caenorhabditis elegans or especially advantageously Phytophtora infestans, Thallasiosira pseudonona and Cryptocodinium cohnii.

Transgenic plants may be obtained by transformation techniques as published, and cited, in: Plant Molecular Biology and Biotechnology (CRC Press, Boca Raton, Fla.), chapter 6/7, pp. 71-119 (1993); F. F. White, Vectors for Gene Transfer in Higher Plants; in: Transgenic Plants, vol. 1, Engineering and Utilization, Ed.: Kung and R. Wu, Academic Press, 1993, 15-38; B. Jenes et al., Techniques for Gene Transfer, in: Transgenic Plants, vol. 1, Engineering and Utilization, Ed.: Kung and R. Wu, Academic Press (1993), 128-143; Potrykus, Annu. Rev. Plant Physiol. Plant Molec. Biol. 42 (1991), 205-225. Preferably, transgenic plants can be obtained by T-DNA-mediated transformation. Such vector systems are, as a rule, characterized in that they contain at least the vir genes, which are required for the Agrobacterium-mediated transformation, and the sequences which delimit the T-DNA (T-DNA border). Suitable vectors are described elsewhere in the specification in detail.

Preferably, a multicellular micro-organism as used herein refers to protists or diatoms. More preferably, it is selected from the group of the families Dinophyceae, Turaniellidae or Oxytrichidae, such as the genera and species: Crypthecodinium cohnii, Phaeodactylum tricornutum, Stylonychia mytilus, Stylonychia pustulate, Stylonychia putrina, Stylonychia notophora, Stylonychia sp., Colpidium campylum or Colpidium sp.

The present invention also relates to a method for expressing a nucleic acid of interest in a host cell comprising

-   -   (a) introducing the polynucleotide or the vector of the present         invention into the host cell, whereby the nucleic acid sequence         of interest will be operatively linked to the expression control         sequence; and     -   (b) expressing the said nucleic acid sequence in said host cell.

The polynucleotide or vector of the present invention can be introduced into the host cell by suitable transfection or transformation techniques as specified elsewhere in this description. The nucleic acid of interest will be expressed in the host cell under suitable conditions. To this end, the host cell will be cultivated under conditions which, in principle, allow for transcription of nucleic acids. Moreover, the host cell, preferably, comprises the exogenously supplied or endogenously present transcription machinery required for expressing a nucleic acid of interest by the expression control sequence. More preferably, the host cell is a plant cell and, most preferably, a seed cell or precursor thereof.

Moreover, the present invention encompasses a method for expressing a nucleic acid of interest in a non-human organism comprising

-   -   (a) introducing the polynucleotide or the vector of the present         invention into the non human organism, whereby the nucleic acid         sequence of interest will be operatively linked to the         expression control sequence; and     -   (b) expressing the said nucleic acid sequence in said non-human         transgenic organism.

The polynucleotide or vector of the present invention can be introduced into the non-human transgenic organism by suitable techniques as specified elsewhere in this description. The non-human transgenic organism, preferably, comprises the exogenously supplied or endogenously present transcription machinery required for expressing a nucleic acid of interest by the expression control sequence. More preferably, the non-human transgenic organism is a plant or seed thereof. It is to be understood that the nucleic acid of interest will be expressed, preferably, seed specific in the said non-human transgenic organism.

Further, the present invention relates to a method for the manufacture of stearidonic acid (SDA) in a plant seed comprising the steps of:

-   a) growing a transgenic plant expressing a polynucleotide encoding a     delta 6 desaturase under the control of the polynucleotide of the     present invention; and -   b) obtaining said SDA from the harvested seeds of the said plant.

The SDA may be manufactured in the form of a triglyceride ester, a phospholipids or as Acyl-CoA bound or free fatty acid comprised by seed oil or seed fatty acid preparations obtainable from the harvested seeds of the grown transgenic plants by standard techniques such as an oil mill or chromatographic extraction and/or purification techniques.

Moreover, how to grow transgenic plants and how to harvest their seeds is well known in the art. How to make transgenic plants expressing a gene of interest such as a delta 6 desaturase under the control of the polynucleotide of the present invention is set forth elsewhere herein.

Surprisingly, the polynucleotides of the present invention were found to influence the ratio of the omega-3 fatty acid stearidonic acid (SDA) to the omega-6 fatty acid gamma linolenic acid (GLA). Accordingly, the seeds of the aforementioned transgenic plants expressing a delta 6 desaturase under the control of the promoter comprised by the polynucleotide of the invention are particularly suitable as a source for SDA or SDA enriched fatty acid preparations such as oil.

Thus, the present invention also pertains to the use of the polynucleotide of the present invention driving expression of a delta 6 desaturase in a transgenic plant for increasing (preferably to a statistically significant extent) the amount of SDA at the expense of GLA in plant seeds of said plants.

Moreover, the present invention pertains to seed oil having the said altered SDA to GLA ratio (i.e. an increased SDA amount at the expense of GLA) obtainable by an oil mill from the harvested seeds of a transgenic plant as set forth above. It will be understood that such an oil will be characterized in addition to the altered SDA to GLA ratio by the presence of remaining DNA contaminations including the polynucleotide of the present invention and/or the delta 6 desaturase encoding polynucleotide.

Nucleic acids encoding suitable delta 6 desaturases are well known in the art and are d6-Desaturases d6Des(Cp) from Ceratodon purpureus (WO2000075341), d6Des(OI) from Ostreococcus lucimarinus (WO2008040787), d6Des(Ot) from Ostreococcus tauri (WO2006069710), d6Des(Pf) from Primula farinosa (WO2003072784), d6Des(Pir)_BO from Pythium irregulare (WO2002026946), d6Des(Pir) from Pythium irregulare (WO2002026946), d6Des(Plu) from Primula luteola (WO2003072784), d6Des(Pp) from Physcomitrella patens (WO200102591), d6Des(Pt) from Phaeodactylum tricornutum (WO2002057465), d6Des(Pv) from Primula vialii (WO2003072784) and d6Des(Tp) from Thalassiosira pseudonana (WO2006069710) and, in particular, those mentioned in the accompanying Examples.

Moreover, a transgenic plant expressing a polynucleotide encoding a delta 6 desaturase under the control of the polynucleotide of the present invention may also comprise further desaturases or elongases of the omega-3 pathway which are required for the synthesis of end products such as eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA).

Thus, a method is provided for increasing stearidonic acid in a plant seed comprising the steps of growing a transgenic plant expressing a polynucleotide encoding a delta 6 desaturase under the control of the polynucleotide of the present invention.

By increasing the omega-3 pathway substrate SDA at the expense of the omega-6 pathway substrate GLA, further fatty acid products of the omega-3 pathway can be produced more efficiently in the aforementioned transgenic plants. Preferably, the desaturases and/or elongases required for the production of a desired fatty acid can also be expressed under the control of a polynucleotide of the present invention. Most preferably, however, it is envisaged that the delta 6 desaturase and the further desaturases and/or elongases are expressed under the control of polynucleotides of the present invention comprising different expression control sequences with respect to each other.

In the following tables 1 to 9, the cis-regulatory elements found in the expression control sequences of the present invention are shown.

TABLE 1 cis-regulatory elements of SEQ ID NO: 1 Seq. Opt. Start End Core Matrix name Family/matrix Further Information thresh. pos. pos. Strand sim. sim. Sequence SEQ_1 P$AHBP/ATHB9.01 HD-ZIP class III protein ATHB9 0.77 7 17 (−) 1.000 0.772 ttgATGAtttc SEQ_1 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain protein 0.82 57 77 (+) 1.000 0.897 aaaaaCCATatcttgaaaacc AGAMOUS-like 2 SEQ_1 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 57 75 (+) 0.750 0.827 aaAAACcatatcttgaaaa sugar-responsive genes SEQ_1 P$TELO/ATPURA.01 Arabidopsis Telo-box interacting protein related 0.85 57 71 (+) 0.750 0.851 aaaaACCAtatcttg to the conserved animal protein Pur- alpha SEQ_1 P$GTBX/GT3A.01 Trihelix DNA-binding factor GT-3a 0.83 86 102 (−) 1.000 0.847 accattGTTActcccct SEQ_1 P$LFYB/LFY.01 Plant specific floral meristem identity gene 0.93 91 103 (−) 0.914 0.936 tACCAttgttact LEAFY (LFY) SEQ_1 P$SEF3/SEF3.01 SEF3, Soybean embryo factor 3 0.87 147 161 (+) 1.000 0.875 acccaACCCaaagag SEQ_1 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 177 191 (+) 1.000 0.943 tcagctaaAATCtaa SEQ_1 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 184 194 (+) 1.000 0.921 aaATCTaagga biosynthesis and in the expression of photo- synthesis-related genes SEQ_1 P$LEGB/RY.01 RY and Sph motifs conserved in seed-specific 0.87 202 228 (−) 1.000 0.936 ggctactcCATGcaatattggatgctc promoters SEQ_1 P$CAAT/CAAT.01 CCAAT-box in plant promoters 0.97 206 214 (+) 1.000 0.983 atCCAAtat SEQ_1 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 230 240 (+) 0.878 0.865 aAAATgatgcg SEQ_1 P$E2FF/E2F.01 E2F class I sites 0.82 234 248 (−) 0.757 0.833 ttgtTTCTcgcatca SEQ_1 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 274 284 (+) 1.000 0.918 cgATCTacaat biosynthesis and in the expression of photo- synthesis-related genes SEQ_1 P$L1BX/PDF2.01 Protodermal factor 2 0.85 278 294 (+) 1.000 0.899 ctacaaTAAAtaccaga SEQ_1 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 287 301 (+) 1.000 0.881 ataccagaAATCtca SEQ_1 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix DNA- 0.85 330 346 (+) 0.843 0.903 catgagGTGAgtctttt binding domain SEQ_1 P$OPAQ/GCN4.01 GCN4, conserved in cereal seed storage 0.81 332 348 (+) 1.000 0.830 tgaggTGAGtctttttt protein gene promoters, similar to yeast GCN4 and vertebrate AP-1 SEQ_1 P$SALT/ALFIN1.02 Zinc-finger protein in alfalfa roots, regulates 0.95 360 374 (−) 1.000 0.977 ggtatgcGGTGtttc salt tolerance SEQ_1 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 367 389 (+) 0.812 0.736 cgcataccagaAACGtaaagaaa SEQ_1 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 375 391 (+) 1.000 0.815 agaaacgtAAAGaaaat protein gene promoters SEQ_1 P$HEAT/HSE.01 Heat shock element 0.81 375 389 (+) 1.000 0.918 agaaacgtaaAGAAa SEQ_1 P$MADS/MADS.01 Binding sites for AP1, AP3-PI and AG dimers 0.75 411 431 (−) 1.000 0.791 ttttcCCATattttttacatt SEQ_1 P$TBPF/TATA.01 Plant TATA box 0.88 464 478 (+) 1.000 0.941 aaaaTATAaaaaaaa SEQ_1 P$GAPB/GAP.01 Cis-element in the GAPDH promoters conferring 0.88 474 488 (+) 0.807 0.895 aaaaATTAaaagaaa light inducibility SEQ_1 P$PSRE/GAAA.01 GAAA motif involved in pollen specific transcriptional 0.83 480 496 (+) 1.000 0.838 taaaaGAAAattttgac activation SEQ_1 P$WBXF/WRKY.01 WRKY plant specific zinc-finger-type factor 0.92 487 503 (+) 1.000 0.920 aaattTTGAcgctgaaa associated with pathogen defence, W box SEQ_1 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 498 514 (+) 0.750 0.801 ctgaaaccGTAAatctt SEQ_1 P$TELO/ATPURA.01 Arabidopsis Telo-box interacting protein related 0.85 499 513 (+) 0.750 0.863 tgaaACCGtaaatct to the conserved animal protein Pur- alpha SEQ_1 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 501 515 (+) 1.000 0.889 aaaccgtaAATCtta SEQ_1 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 517 533 (+) 0.750 0.851 aatcaaCAAAtgcataa meristem layer 1) SEQ_1 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 525 541 (+) 1.000 0.954 aatgcaTAAAtgcaaag meristem layer 1) SEQ_1 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 530 546 (+) 1.000 0.773 ataaatgcAAAGttatt protein gene promoters SEQ_1 P$AHBP/BLR.01 Transcriptional repressor BELLRINGER 0.90 538 548 (+) 0.826 0.936 aaaGTTAttga SEQ_1 P$TBPF/TATA.01 Plant TATA box 0.88 557 571 (−) 1.000 0.964 ctaaTATAaaaatat SEQ_1 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 583 599 (−) 1.000 0.934 tagtatttGTTAgcagt SEQ_1 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds 0.87 593 605 (+) 1.000 0.902 aaTACTatacaga to the SP8a (ACTGTGTA) and SP8b (TAC- TATT) sequences of sporamin and beta- amylase genes SEQ_1 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 600 612 (−) 1.000 0.955 tgttTTGTctgta SEQ_1 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 605 623 (+) 0.750 0.836 acAAAAcacattattaaaa sugar-responsive genes SEQ_1 P$AHBP/HAHB4.01 Sunflower homeodomain leucine-zipper protein 0.87 611 621 (+) 1.000 0.909 cacattATTAa Hahb-4 SEQ_1 P$GTBX/SBF1.01 SBF-1 0.87 611 627 (+) 1.000 0.886 cacattaTTAAaaaaac SEQ_1 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 627 639 (−) 1.000 0.965 tattTTGTctttg SEQ_1 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix DNA- 0.85 636 652 (−) 1.000 0.858 tgatatGTTAatatatt binding domain SEQ_1 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds 0.87 649 661 (+) 0.814 0.901 atCACTattacta to the SP8a (ACTGTGTA) and SP8b (TAC- TATT) sequences of sporamin and beta- amylase genes SEQ_1 P$L1BX/ATML1.02 Arabidopsis thaliana meristem layer 1 0.76 669 685 (+) 0.890 0.762 acaCAATaaaaacacca SEQ_1 P$CARM/CARICH.01 CA-rich element 0.78 672 690 (+) 1.000 0.815 caataaaAACAccaaataa SEQ_1 P$DOFF/PBF.01 PBF (MPBF) 0.97 698 714 (+) 1.000 0.990 aacaaataAAAGtgatc SEQ_1 P$OCSE/OCSL.01 OCS-like elements 0.69 701 721 (+) 0.807 0.729 aaataaaagtgatcACATaat SEQ_1 P$OCSE/OCSL.01 OCS-like elements 0.69 704 724 (−) 0.769 0.713 gtaattatgtgatcACTTtta SEQ_1 P$AHBP/HAHB4.01 Sunflower homeodomain leucine-zipper protein 0.87 714 724 (+) 1.000 0.902 cacataATTAc Hahb-4 SEQ_1 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix DNA- 0.85 714 730 (−) 0.968 0.867 gataatGTAAttatgtg binding domain SEQ_1 P$IBOX/GATA.01 Class I GATA factors 0.93 719 735 (−) 1.000 0.942 ttttgGATAatgtaatt SEQ_1 P$MYBS/MYBST1.01 MybSt1 (Myb Solanum tuberosum 1) with a 0.90 722 738 (+) 1.000 0.962 tacattATCCaaaaaat single myb repeat SEQ_1 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 733 751 (+) 1.000 0.908 aaAAATcatacttttaaca sugar-responsive genes SEQ_1 P$GTBX/SBF1.01 SBF-1 0.87 740 756 (−) 1.000 0.967 attgttgTTAAaagtat SEQ_1 P$MYBS/TAMYB80.01 MYB protein from wheat 0.83 759 775 (+) 1.000 0.857 aacaATATtccgcgcga SEQ_1 P$CGCG/OSCBT.01 Oryza sativa CaM-binding transcription factor 0.78 768 784 (−) 1.000 0.781 gtcCGCGcttcgcgcgg SEQ_1 P$NCS3/NCS3.01 Nodulin consensus sequence 3 0.89 782 792 (+) 1.000 0.913 gaCACCcccct SEQ_1 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription factors 0.81 802 818 (−) 1.000 0.829 catacaACATgactaca that belong to the group of Opaque-2 like proteins SEQ_1 P$TEFB/TEF1.01 TEF cis acting elements in both RNA polymerase 0.76 842 862 (−) 0.838 0.843 gcATGGgaaatcaggtccatc II-dependent promoters and rDNA spacer sequences SEQ_1 P$EINL/TEIL.01 TEIL (tobacco EIN3-like) 0.92 843 851 (+) 0.863 0.966 aTGGAcctg SEQ_1 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 847 873 (−) 0.750 0.592 acgactaCTATgcatgggaaatcaggt element about 100 bp upstream of the TSS in legumin genes SEQ_1 P$LEGB/RY.01 RY and Sph motifs conserved in seed-specific 0.87 850 876 (+) 1.000 0.944 tgatttccCATGcatagtagtcgtcat promoters SEQ_1 P$MADS/AGL15.01 AGL15, Arabidopsis MADS-domain protein 0.79 851 871 (−) 1.000 0.850 gacTACTatgcatgggaaatc AGAMOUS-like 15 SEQ_1 P$MADS/AGL3.01 AGL3, MADS Box protein 0.83 852 872 (+) 1.000 0.864 atttcCCATgcatagtagtcg SEQ_1 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds 0.87 858 870 (−) 1.000 0.922 acTACTatgcatg to the SP8a (ACTGTGTA) and SP8b (TAC- TATT) sequences of sporamin and beta- amylase genes SEQ_1 P$GBOX/TGA1.01 Arabidopsis leucine zipper protein TGA1 0.90 861 881 (−) 1.000 0.900 ccgagaTGACgactactatgc SEQ_1 P$TELO/ATPURA.01 Arabidopsis Telo-box interacting protein related 0.85 873 887 (−) 1.000 0.860 ataaACCCgagatga to the conserved animal protein Pur- alpha SEQ_1 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription factors 0.81 883 899 (−) 0.829 0.887 gatataACTTgaataaa that belong to the group of Opaque-2 like proteins SEQ_1 P$DOFF/DOF1.01 Dof1/MNB1a - single zinc finger transcription 0.98 912 928 (−) 1.000 0.993 ttcagattAAAGaacgt factor SEQ_1 P$MADS/AGL15.01 AGL15, Arabidopsis MADS-domain protein 0.79 912 932 (+) 0.925 0.793 acgTTCTttaatctgaaccct AGAMOUS-like 15 SEQ_1 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 915 925 (+) 1.000 0.963 ttcttTAATct SEQ_1 P$TELO/ATPURA.01 Arabidopsis Telo-box interacting protein related 0.85 924 938 (+) 1.000 0.876 ctgaACCCtatcacc to the conserved animal protein Pur- alpha SEQ_1 P$STKM/STK.01 Storekeeper (STK), plant specific DNA binding 0.85 937 951 (−) 1.000 0.877 tccTAAAtaaatcgg protein important for tuber-specific and sucrose- inducible gene expression SEQ_1 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix DNA- 0.85 968 984 (−) 0.968 0.858 aaagtgGTAAtttttgt binding domain SEQ_1 P$DOFF/PBF.01 PBF (MPBF) 0.97 976 992 (−) 1.000 0.988 gagacagaAAAGtggta SEQ_1 P$GTBX/SBF1.01 SBF-1 0.87 999 1015 (+) 1.000 0.907 ctcgtttTTAAtttggt SEQ_1 P$MIIG/MYBC1.01 Maize C1 myb-domain protein 0.92 1009 1023 (+) 1.000 0.942 atttgGTAGtttcag SEQ_1 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds 0.87 1048 1060 (−) 0.814 0.872 aaCACTatgaaaa to the SP8a (ACTGTGTA) and SP8b (TAC- TATT) sequences of sporamin and beta- amylase genes SEQ_1 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription factors 0.81 1056 1072 (+) 1.000 0.891 gtgttaACATgtttaag that belong to the group of Opaque-2 like proteins SEQ_1 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 1095 1105 (−) 1.000 0.858 atATCTatgtt biosynthesis and in the expression of photo- synthesis-related genes SEQ_1 P$MYBL/NTMYBAS1.01 Anther-specific myb gene from tobacco 0.96 1125 1141 (+) 1.000 0.967 tagtggtgGTTAacaaa SEQ_1 P$GTBX/SBF1.01 SBF-1 0.87 1127 1143 (+) 1.000 0.894 gtggtggTTAAcaaaag SEQ_1 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 1130 1146 (−) 1.000 0.918 ggccttttGTTAaccac SEQ_1 P$GBOX/BZIP911.01 bZIP transcription factor from Antirrhinum 0.77 1138 1158 (−) 0.750 0.781 ataagtTGAAatggccttttg majus SEQ_1 P$OPAQ/O2.01 Opaque-2 regulatory protein 0.87 1141 1157 (+) 0.852 0.882 aaggccattTCAActta SEQ_1 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 1163 1173 (+) 1.000 0.909 tAAAAgataga SEQ_1 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 1176 1194 (+) 0.750 0.815 gcAAAGcattgttgataaa sugar-responsive genes SEQ_1 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription factor 0.99 1184 1200 (+) 1.000 0.994 ttgttgatAAAGcctct SEQ_1 P$GAGA/BPC.01 Basic pentacysteine proteins 1.00 1184 1208 (−) 1.000 1.000 ataaagAGAGaggctttatcaacaa SEQ_1 P$IBOX/GATA.01 Class I GATA factors 0.93 1184 1200 (+) 1.000 0.938 ttgttGATAaagcctct SEQ_1 P$MYBS/ZMMRP1.01 Zea mays MYB-related protein 1 (transfer cell 0.79 1199 1215 (+) 0.777 0.841 ctctcttTATAtaaaga specific) SEQ_1 P$TBPF/TATA.01 Plant TATA box 0.88 1199 1213 (−) 1.000 0.958 tttaTATAaagagag SEQ_1 P$TBPF/TATA.02 Plant TATA box 0.90 1201 1215 (−) 1.000 0.917 tcttTATAtaaagag SEQ_1 P$TBPF/TATA.02 Plant TATA box 0.90 1202 1216 (+) 1.000 0.917 tcttTATAtaaagag SEQ_1 P$TBPF/TATA.01 Plant TATA box 0.88 1204 1218 (+) 1.000 0.934 tttaTATAaagaggg SEQ_1 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription factor 0.99 1215 1231 (−) 1.000 0.995 aaggcgctAAAGcccct SEQ_1 P$WBXF/WRKY.01 WRKY plant specific zinc-finger-type factor 0.92 1236 1252 (+) 1.000 0.975 atgctTTGActttacct associated with pathogen defence, W box SEQ_1 P$TELO/RPBX.01 Ribosomal protein box, appears unique to 0.84 1260 1274 (−) 1.000 0.842 cgaaaCCCTtcactt plant RP genes and genes associated with gene expression SEQ_1 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 1317 1333 (+) 0.750 0.830 caagaaTCAAtgtaagc meristem layer 1) SEQ_1 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 1332 1346 (−) 1.000 0.879 caaactatAATCtgc SEQ_1 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 1338 1354 (+) 0.817 0.929 tatagtTTGTtagtttt SEQ_1 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 1342 1358 (+) 1.000 0.818 gtttgtTAGTttttcag SEQ_1 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 1382 1398 (−) 1.000 0.818 aggcgtTAGTtcagaaa SEQ_1 P$MYBL/NTMYBAS1.01 Anther-specific myb gene from tobacco 0.96 1386 1402 (−) 1.000 0.967 tcaaaggcGTTAgttca SEQ_1 P$MADS/SQUA.01 MADS-box protein SQUAMOSA 0.90 1400 1420 (−) 1.000 0.902 attgaccATTTttttctttca SEQ_1 P$WBXF/ERE.01 Elicitor response element 0.89 1408 1424 (−) 1.000 0.973 gtcaatTGACcattttt SEQ_1 P$GTBX/GT3A.01 Trihelix DNA-binding factor GT-3a 0.83 1425 1441 (−) 1.000 0.897 ttagtgGTTAcaatggc SEQ_1 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix DNA- 0.85 1432 1448 (−) 1.000 0.882 catgtgGTTAgtggtta binding domain SEQ_1 P$MIIG/PALBOXP.01 Putative cis-acting element in various PAL and 0.81 1433 1447 (−) 0.936 0.820 atGTGGttagtggtt 4CL gene promoters SEQ_1 P$LEGB/RY.01 RY and Sph motifs conserved in seed-specific 0.87 1445 1471 (−) 1.000 0.944 tgtgtttgCATGcatagccagtgcatg promoters SEQ_1 P$LEGB/RY.01 RY and Sph motifs conserved in seed-specific 0.87 1452 1478 (+) 1.000 0.909 tggctatgCATGcaaacacaatgagat promoters SEQ_1 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds 0.87 1488 1500 (−) 1.000 0.871 ttTACTcttaggc to the SP8a (ACTGTGTA) and SP8b (TAC- TATT) sequences of sporamin and beta- amylase genes SEQ_1 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 1493 1509 (−) 0.778 0.838 caaagtTGGTttactct SEQ_1 P$MADS/AGL1.01 AGL1, Arabidopsis MADS-domain protein 0.84 1495 1515 (−) 0.975 0.840 ggaTTCCaaagttggtttact AGAMOUS-like 1 SEQ_1 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain protein 0.82 1496 1516 (+) 0.968 0.845 gtaaaCCAActttggaatccc AGAMOUS-like 2 SEQ_1 P$STKM/STK.01 Storekeeper (STK), plant specific DNA binding 0.85 1513 1527 (+) 0.833 0.873 tccCAAAaaattata protein important for tuber-specific and sucrose- inducible gene expression SEQ_1 P$ERSE/ERSE_I.01 ERSE I (ER stress-response element I)-like 0.79 1515 1533 (+) 0.750 0.803 ccaaaaaattatagcCATG motif SEQ_1 P$GBOX/EMBP1.01 bZIP transcription factor implicated in ABA 0.84 1522 1542 (−) 0.750 0.854 agaacgacACATggctataat induced gene expression SEQ_1 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 1522 1548 (+) 1.000 0.609 attatagCCATgtgtcgttcttgatga element about 100 bp upstream of the TSS in legumin genes SEQ_1 P$ABRE/ABF1.01 ABA (abscisic acid) inducible transcriptional 0.79 1525 1541 (−) 1.000 0.853 gaacgACACatggctat activator SEQ_1 P$OCSE/OCSL.01 OCS-like elements 0.69 1549 1569 (−) 0.807 0.693 aaattttattggaaACGAatt SEQ_1 P$GTBX/SBF1.01 SBF-1 0.87 1562 1578 (−) 0.826 0.872 tttgtttCTAAatttta SEQ_1 P$MIIG/PALBOXP.01 Putative cis-acting element in various PAL and 0.81 1570 1584 (−) 0.936 0.819 ttGTGGtttgtttct 4CL gene promoters SEQ_1 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds 0.87 1586 1598 (−) 1.000 0.881 atTACTttgtatt to the SP8a (ACTGTGTA) and SP8b (TAC- TATT) sequences of sporamin and beta- amylase genes SEQ_1 P$OCSE/OCSL.01 OCS-like elements 0.69 1589 1609 (−) 0.769 0.716 taagttaaaaaattACTTtgt SEQ_1 P$AHBP/BLR.01 Transcriptional repressor BELLRINGER 0.90 1591 1601 (−) 1.000 0.976 aaaATTActtt SEQ_1 P$STKM/STK.01 Storekeeper (STK), plant specific DNA binding 0.85 1593 1607 (−) 1.000 0.901 agtTAAAaaattact protein important for tuber-specific and sucrose- inducible gene expression SEQ_1 P$MADS/MADS.01 Binding sites for AP1, AP3-PI and AG dimers 0.75 1599 1619 (−) 1.000 0.777 tttccCCATttaagttaaaaa SEQ_1 P$L1BX/PDF2.01 Protodermal factor 2 0.85 1602 1618 (+) 1.000 0.864 ttaactTAAAtggggaa SEQ_1 P$IBOX/IBOX.01 I-Box in rbcS genes and other light regulated 0.81 1640 1656 (+) 1.000 0.824 aaagaGATAgggcttaa genes SEQ_1 P$TELO/RPBX.01 Ribosomal protein box, appears unique to 0.84 1642 1656 (−) 1.000 0.886 ttaagCCCTatctct plant RP genes and genes associated with gene expression SEQ_1 P$DOFF/DOF2.01 Dof2 - single zinc finger transcription factor 0.98 1647 1663 (+) 1.000 0.994 tagggcttAAAGcagca SEQ_1 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 1673 1695 (−) 0.812 0.695 aacgaaaacgaAACGtatcacag SEQ_1 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 1689 1705 (+) 1.000 0.963 tttcgtttGTTAtcaca SEQ_1 P$IBOX/GATA.01 Class I GATA factors 0.93 1691 1707 (−) 1.000 0.931 attgtGATAacaaacga SEQ_1 P$GTBX/S1F.01 S1F, site 1 binding factor of spinach rps1 0.79 1707 1723 (+) 1.000 0.851 tttcATGGactatatac promoter SEQ_1 P$TBPF/TATA.02 Plant TATA box 0.90 1713 1727 (+) 1.000 0.901 ggacTATAtacattt SEQ_1 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 1718 1734 (−) 1.000 0.875 ctaagcTAAAtgtatat meristem layer 1) SEQ_1 P$PSRE/GAAA.01 GAAA motif involved in pollen specific transcriptional 0.83 1739 1755 (+) 1.000 0.847 caaaaGAAAccatctac activation SEQ_1 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 1748 1758 (+) 1.000 0.854 ccATCTacttg biosynthesis and in the expression of photo- synthesis-related genes SEQ_1 P$AHBP/ATHB1.01 Arabidopsis thaliana homeo box protein 1 0.90 1768 1778 (−) 1.000 0.990 ggaATTAttgt SEQ_1 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 1768 1778 (+) 0.829 0.940 acaATAAttcc

TABLE 2 cis-regulatory elements of SEQ ID NO: 6 Seq. Opt. Start End Core Matrix name Family/matrix Further Information thresh. pos. pos. Strand sim. sim. Sequence SEQ_6 P$SEF3/SEF3.01 SEF3, Soybean embryo factor 3 0.87 5 19 (+) 1.000 0.921 ctataACCCaaccca SEQ_6 P$SEF3/SEF3.01 SEF3, Soybean embryo factor 3 0.87 10 24 (+) 1.000 0.891 acccaACCCaaaaca SEQ_6 P$GTBX/SBF1.01 SBF-1 0.87 22 38 (−) 1000 0.880 aggatacTTAAacttgt SEQ_6 P$MYBL/ATMYB77.01 R2R3-type myb-like transcription factor (I-type 0.87 35 51 (−) 1000 0.892 ttttgtCGGTttcagga binding site) SEQ_6 P$DREB/CRT_DRE.01 C-repeat/dehydration response element 0.89 38 52 (+) 1.000 0.902 tgaaaCCGAcaaaag SEQ_6 P$DOFF/PBF.01 PBF (MPBF) 0.97 41 57 (+) 1000 0.986 aaccgacaAAAGagaat SEQ_6 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 41 53 (−) 1.000 0.987 tcttTTGTcggtt SEQ_6 P$EPFF/ZPT22.01 Member of the EPF family of zinc finger transcription 0.75 50 72 (−) 1.000 0.770 agtcaaaaagaCAGTattctctt factors SEQ_6 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds to 0.87 55 67 (+) 1.000 0.881 aaTACTgtctttt the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_6 P$NCS2/NCS2.01 Nodulin consensus sequence 2 0.79 57 71 (+) 0.750 0.795 tactgtCTTTttgac SEQ_6 P$WBXF/WRKY.01 WRKY plant specific zinc-finger-type factor associated 0.92 62 78 (+) 1.000 0.965 tctttTTGActttcctg with pathogen defence, W box SEQ_6 P$HEAT/HSE.01 Heat shock element 0.81 72 86 (−) 0.826 0.819 agattattcaGGAAa SEQ_6 P$AHBP/BLR.01 Transcriptional repressor BELLRINGER 0.90 77 87 (−) 1000 0.901 aagATTAttca SEQ_6 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 80 96 (−) 1.000 0.769 tattttttAAAGattat protein gene promoters SEQ_6 P$GTBX/SBF1.01 SBF-1 0.87 80 96 (+) 1.000 0.892 ataatctTTAAaaaata SEQ_6 P$GAGA/BPC.01 Basic pentacysteine proteins 1.00 88 112 (−) 1.000 1000 ttccagAGAGaactgatatttttta SEQ_6 P$PSRE/GAAA.01 GAAA motif involved in pollen specific transcriptional 0.83 105 121 (+) 1000 0.878 ctctgGAAAtagtaaag activation SEQ_6 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds to 0.87 108 120 (−) 1.000 0.943 ttTACTatttcca the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_6 P$OCSE/OCSL.01 OCS-like elements 0.69 116 136 (−) 0.769 0.700 ttgcatcagttcttACTTtac SEQ_6 P$HOCT/HOCT.01 Octamer motif found in plant histone H3 and H4 0.76 145 161 (+) 0.750 0.762 atcaccgATCTacgaag genes SEQ_6 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol bio- 0.85 150 160 (+) 1.000 0.864 cgATCTacgaa synthesis and in the expression of photosynthesis- related genes SEQ_6 P$MSAE/MSA.01 M-phase-specific activators (NtmybA1, NtmybA2, 0.80 174 188 (−) 1.000 0.834 aaaaaAACGggtgga NtmybB) SEQ_6 P$AHBP/HAHB4.01 Sunflower homeodomain leucine-zipper protein 0.87 187 197 (+) 1.000 0.934 ttgatcATTAt Hahb-4 SEQ_6 P$CGCG/ATSR1.01 Arabidopsis thaliana signal-responsive gene1, 0.84 194 210 (−) 1.000 0.912 aagCGCGtacgatataa Ca2+/calmodulin binding protein homolog to NtER1 (tobacco early ethylene-responsive gene) SEQ_6 P$OCSE/OCSL.01 OCS-like elements 0.69 196 216 (−) 0.807 0.709 gtttttaagcgcgtACGAtat SEQ_6 P$GTBX/SBF1.01 SBF-1 0.87 202 218 (+) 1.000 0.875 tacgcgcTTAAaaacct SEQ_6 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 217 233 (−) 0.778 0.781 tttagtTCGTaaaaaag SEQ_6 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 245 261 (−) 1000 0.831 ttatctgaAAAGtaaaa protein gene promoters SEQ_6 P$IBOX/GATA.01 Class I GATA factors 0.93 252 268 (+) 1000 0.933 tttcaGATAatgttgca SEQ_6 P$AGP1/AGP1.01 AG-motif binding protein 1 0.91 284 294 (−) 1.000 0.988 acaGATCtatc SEQ_6 P$AGP1/AGP1.01 AG-motif binding protein 1 0.91 285 295 (+) 1000 0.920 ataGATCtgtt SEQ_6 P$GARP/ARR10.01 Type-B response regulator (ARR10), member of 0.97 287 295 (+) 1.000 0.970 AGATctgtt the GARP-family of plant myb-related DNA binding motifs SEQ_6 P$CAAT/CAAT.01 CCAAT-box in plant promoters 0.97 298 306 (+) 1.000 0.973 ttCCAAtga SEQ_6 P$LFYB/LFY.01 Plant specific floral meristem identity gene LEAFY 0.93 298 310 (+) 1000 0.930 tTCCAatgagaat (LFY) SEQ_6 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 300 314 (+) 1.000 0.864 ccaatgagAATCtgt SEQ_6 P$GARP/ARR10.01 Type-B response regulator (ARR10), member of 0.97 304 312 (−) 1.000 0.971 AGATtctca the GARP-family of plant myb-related DNA binding motifs SEQ_6 P$RAV5/RAV1-5.01 5′-part of bipartite RAV1 binding site, interacting 0.96 308 318 (−) 1.000 0.960 aacAACAgatt with AP2 domain SEQ_6 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 339 351 (−) 1.000 0.934 tattTTGTcagat SEQ_6 P$CAAT/CAAT.02 CCAAT-box in plant promoters 1.00 400 408 (−) 1000 1000 gtcCAATta SEQ_6 P$WBXF/WRKY.01 WRKY plant specific zinc-finger-type factor associated 0.92 405 421 (−) 1.000 0.957 cacatTTGActatgtcc with pathogen defence, W box SEQ_6 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), AtMYC2 0.95 407 425 (−) 1.000 0.953 ccaacACATttgactatgt (rd22BP1) SEQ_6 P$CARM/CARICH.01 CA-rich element 0.78 412 430 (−) 1.000 0.785 aaagtccAACAcatttgac SEQ_6 P$GTBX/SBF1.01 SBF-1 0.87 424 440 (−) 1.000 0.875 tcggaaaTTAAaagtcc SEQ_6 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 439 457 (+) 1000 0.913 gaAAATcattaaaaacaat sugar-responsive genes SEQ_6 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 440 450 (−) 0.829 0.902 ttaATGAtttt SEQ_6 P$AHBP/HAHB4.01 Sunflower homeodomain leucine-zipper protein 0.87 440 450 (+) 1.000 0.967 aaaatcATTAa Hahb-4 SEQ_6 P$GTBX/SBF1.01 SBF-1 0.87 440 456 (+) 1.000 0.886 aaaatcaTTAAaaacaa SEQ_6 P$GTBX/SBF1.01 SBF-1 0.87 441 457 (−) 1000 0.888 attgtttTTAAtgattt SEQ_6 P$L1BX/ATML1.02 Arabidopsis thaliana meristem layer 1 0.76 442 458 (+) 1000 0.789 aatCATTaaaaacaatt SEQ_6 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 444 454 (−) 1.000 1000 gttttTAATga SEQ_6 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 448 464 (+) 0.750 0.812 taaaaacaATTAaaaaa SEQ_6 P$TEFB/TEF1.01 TEF cis acting elements in both RNA polymerase 0.76 467 487 (+) 0.838 0.839 taATGGagatttttgtaatta II-dependent promoters and rDNA spacer sequences SEQ_6 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 471 485 (−) 1.000 0.948 attacaaaAATCtcc SEQ_6 P$GTBX/SBF1.01 SBF-1 0.87 478 494 (+) 1.000 0.905 tttgtaaTTAAttggaa SEQ_6 P$CAAT/CAAT.02 CCAAT-box in plant promoters 1.00 486 494 (−) 1.000 1000 ttcCAATta SEQ_6 P$MADS/AGL15.01 AGL15, Arabidopsis MADS-domain protein 0.79 509 529 (−) 1.000 0.886 ctaTACTattaaagggaaaga AGAMOUS-like 15 SEQ_6 P$MADS/AGL3.02 AGL3, MADS Box protein 0.80 510 530 (+) 0.790 0.859 ctttcCCTTtaatagtataga SEQ_6 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds to 0.87 516 528 (−) 1.000 0.956 taTACTattaaag the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_6 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol bio- 0.85 523 533 (−) 1.000 0.902 atATCTatact synthesis and in the expression of photosynthesis- related genes SEQ_6 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 573 589 (−) 0.750 0.844 taatttTAACtgcaact meristem layer 1) SEQ_6 P$GTBX/SBF1.01 SBF-1 0.87 574 590 (+) 1000 0.954 gttgcagTTAAaattac SEQ_6 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 577 599 (+) 1000 0.707 gcagttaaaatTACGaatcatgg SEQ_6 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain protein AGA- 0.82 584 604 (−) 1.000 0.820 ggagcCCATgattcgtaattt MOUS-like 2 SEQ_6 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol bio- 0.85 604 614 (+) 1.000 0.897 ctATCTatatt synthesis and in the expression of photosynthesis- related genes SEQ_6 P$MYBS/ZMMRP1.01 Zea mays MYB-related protein 1 (transfer cell 0.79 604 620 (+) 0.777 0.905 ctatctaTATTttacat specific) SEQ_6 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 608 624 (−) 0.761 0.835 tgtgatgtAAAAtatag protein gene promoters SEQ_6 P$DOFF/DOF2.01 Dof2 - single zinc finger transcription factor 0.98 619 635 (+) 1.000 0.995 atcacaatAAAGctata SEQ_6 P$TBPF/TATA.02 Plant TATA box 0.90 628 642 (+) 1.000 0.903 aagcTATAtatcatt SEQ_6 P$LFYB/LFY.01 Plant specific floral meristem identity gene LEAFY 0.93 634 646 (−) 0.914 0.936 cACCAatgatata (LFY) SEQ_6 P$CAAT/CAAT.01 CCAAT-box in plant promoters 0.97 638 646 (−) 1.000 0.988 caCCAAtga SEQ_6 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 657 669 (−) 1000 0.921 gggtTTGTcttca SEQ_6 P$TELO/RPBX.01 Ribosomal protein box, appears unique to plant 0.84 662 676 (+) 1.000 0.981 acaaaCCCTaaactc RP genes and genes associated with gene expression SEQ_6 P$AHBP/HAHB4.01 Sunflower homeodomain leucine-zipper protein 0.87 684 694 (−) 1.000 0.923 cttattATTAg Hahb-4 SEQ_6 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), AtMYC2 0.95 694 712 (+) 0.954 0.972 gccaaACACttgattccaa (rd22BP1) SEQ_6 P$MADS/SQUA.01 MADS-box protein SQUAMOSA 0.90 711 731 (−) 1000 0.906 ggtcgctATTTgtttctgttt SEQ_6 P$IBOX/GATA.01 Class I GATA factors 0.93 733 749 (+) 1.000 0.958 tgcacGATAatagatag SEQ_6 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol bio- 0.85 739 749 (−) 1.000 0.853 ctATCTattat synthesis and in the expression of photosynthesis- related genes SEQ_6 P$HMGF/HMG_IY.02 High mobility group I/Y-like protein isolated from 1.00 755 769 (−) 1.000 1000 tattTATTtttcaaa pea SEQ_6 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 774 788 (−) 1000 0.864 aaccaagaAATCtga SEQ_6 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds to 0.87 787 799 (−) 1.000 0.909 ttTACTgtttaaa the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_6 P$DOFF/DOF2.01 Dof2 - single zinc finger transcription factor 0.98 790 806 (+) 1.000 0.981 aaacagtaAAAGctaat SEQ_6 P$L1BX/ATML1.02 Arabidopsis thaliana meristem layer 1 0.76 790 806 (+) 0.808 0.780 aaaCAGTaaaagctaat SEQ_6 P$CARM/CARICH.01 CA-rich element 0.78 800 818 (−) 1.000 0.836 tttttgaAACAcattagct SEQ_6 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 823 839 (+) 0.750 0.818 aaaaaacaGTAAaagct SEQ_6 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds to 0.87 823 835 (−) 1000 0.905 ttTACTgtttttt the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_6 P$DOFF/DOF2.01 Dof2 - single zinc finger transcription factor 0.98 826 842 (+) 1000 0.981 aaacagtaAAAGctaat SEQ_6 P$L1BX/ATML1.02 Arabidopsis thaliana meristem layer 1 0.76 826 842 (+) 0.808 0.780 aaaCAGTaaaagctaat SEQ_6 P$GAPB/GAP.01 Cis-element in the GAPDH promoters conferring 0.88 843 857 (+) 1000 0.984 acacATGAagacaag light inducibility SEQ_6 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol bio- 0.85 863 873 (−) 1.000 0.912 aaATCTataag synthesis and in the expression of photosynthesis- related genes SEQ_6 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 866 880 (−) 1.000 0.885 gtgggtaaAATCtat SEQ_6 P$GTBX/SBF1.01 SBF-1 0.87 867 883 (−) 0.782 0.889 tttgtggGTAAaatcta SEQ_6 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 877 893 (−) 0.817 0.846 acaagtTTGTtttgtgg SEQ_6 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain protein AGA- 0.82 906 926 (−) 0.968 0.856 agaagCCAAcattggcaacga MOUS-like 2 SEQ_6 P$MADS/AG.01 Agamous, required for normal flower development, 0.80 907 927 (+) 0.902 0.806 cgtTGCCaatgttggcttctt similarity to SRF (human) and MCM (yeast) proteins SEQ_6 P$LFYB/LFY.01 Plant specific floral meristem identity gene LEAFY 0.93 910 922 (+) 0.885 0.938 tGCCAatgttggc (LFY) SEQ_6 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 920 936 (−) 1.000 0.801 tgtggtggAAAGaagcc protein gene promoters SEQ_6 P$SALT/ALFIN1.01 Zinc-finger protein in alfalfa roots, regulates salt 0.93 926 940 (−) 1.000 0.986 tttgtGTGGtggaaa tolerance SEQ_6 P$TEFB/TEF1.01 TEF cis acting elements in both RNA polymerase 0.76 931 951 (−) 0.838 0.781 taACGGtcatatttgtgtggt II-dependent promoters and rDNA spacer sequences SEQ_6 P$WBXF/ERE.01 Elicitor response element 0.89 937 953 (+) 1.000 0.897 caaataTGACcgttaag SEQ_6 P$MYBL/ATMYB77.01 R2R3-type myb-like transcription factor (I-type 0.87 940 956 (+) 0.857 0.916 atatgaCCGTtaagact binding site) SEQ_6 P$MSAE/MSA.01 M-phase-specific activators (NtmybA1, NtmybA2, 0.80 941 955 (−) 1.000 0.889 gtcttAACGgtcata NtmybB) SEQ_6 P$AHBP/HAHB4.01 Sunflower homeodomain leucine-zipper protein 0.87 968 978 (+) 1000 0.916 tttataATTAc Hahb-4 SEQ_6 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix DNA- 0.85 968 984 (−) 0.968 0.859 catgtaGTAAttataaa binding domain SEQ_6 P$LEGB/RY.01 RY and Sph motifs conserved in seed-specific 0.87 970 996 (−) 1.000 0.952 attttataCATGcatgtagtaattata promoters SEQ_6 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 973 989 (+) 0.750 0.846 aattacTACAtgcatgt meristem layer 1) SEQ_6 P$LEGB/RY.01 RY and Sph motifs conserved in seed-specific 0.87 973 999 (+) 1.000 0.952 aattactaCATGcatgtataaaatcta promoters SEQ_6 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 980 996 (−) 0.750 0.855 attttaTACAtgcatgt meristem layer 1) SEQ_6 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 986 1000 (+) 1.000 0.922 atgtataaAATCtat SEQ_6 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol bio- 0.85 993 1003 (+) 1.000 0.897 aaATCTataga synthesis and in the expression of photosynthesis- related genes SEQ_6 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol bio- 0.85 996 1006 (−) 1000 0.877 cgATCTataga synthesis and in the expression of photosynthesis- related genes

TABLE 3 cis-regulatory elements of SEQ ID NO: 9 Seq. Opt. Start End Core Matrix name Family/matrix Further Information thresh. pos. pos. Strand sim. sim. Sequence SEQ_9 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 17 35 (+) 0.750 0.859 caAATTcaggtagcttaag sugar-responsive genes SEQ_9 P$MIIG/MYBC1.01 Maize C1 myb-domain protein 0.92 21 35 (+) 1.000 0.928 ttcagGTAGcttaag SEQ_9 P$MADS/AGL3.01 AGL3, MADS Box protein 0.83 30 50 (−) 0.973 0.858 agccaCCAAttagagcttaag SEQ_9 P$CAAT/CAAT.01 CCAAT-box in plant promoters 0.97 39 47 (−) 1.000 0.981 caCCAAtta SEQ_9 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription factor 0.99 44 60 (−) 1.000 0.995 tattacctAAAGccacc SEQ_9 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 69 85 (+) 0.817 0.794 ctcagtTTGTaaatgta SEQ_9 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 72 88 (+) 1.000 0.925 agtttgTAAAtgtagtt meristem layer 1) SEQ_9 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 78 94 (+) 1.000 0.794 taaatgTAGTtaaaact SEQ_9 P$GTBX/SBF1.01 SBF-1 0.87 80 96 (+) 1.000 0.929 aatgtagTTAAaacttt SEQ_9 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 88 98 (−) 1.000 0.850 cAAAAgtttta SEQ_9 P$SALT/ALFIN1.01 Zinc-finger protein in alfalfa roots, regulates salt 0.93 93 107 (+) 1.000 0.948 cttttGTGGtgtaaa tolerance SEQ_9 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 97 113 (+) 0.776 0.793 tgtggtgtAAATcatgt protein gene promoters SEQ_9 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix DNA-binding 0.85 97 113 (+) 0.968 0.857 tgtggtGTAAatcatgt domain SEQ_9 P$CAAT/CAAT.01 CCAAT-box in plant promoters 0.97 121 129 (−) 1.000 0.979 aaCCAAtcg SEQ_9 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix DNA-binding 0.85 121 137 (+) 1.000 0.866 cgattgGTTAataaaaa domain SEQ_9 P$SEF4/SEF4.01 Soybean embryo factor 4 0.98 129 139 (−) 1.000 0.985 acTTTTtatta SEQ_9 P$OPAQ/GCN4.01 GCN4, conserved in cereal seed storage protein 0.81 141 157 (+) 1.000 0.813 gttgaTGAGttaaaaaa gene promoters, similar to yeast GCN4 and vertebrate AP-1 SEQ_9 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 158 174 (+) 1.000 0.805 caaaatTAGTtgcagtt SEQ_9 P$AHBP/BLR.01 Transcriptional repressor BELLRINGER 0.90 159 169 (+) 1.000 0.981 aaaATTAgttg SEQ_9 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 165 181 (−) 0.750 0.844 taatttTAACtgcaact meristem layer 1) SEQ_9 P$GTBX/SBF1.01 SBF-1 0.87 166 182 (+) 1.000 0.954 gttgcagTTAAaattac SEQ_9 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 169 191 (+) 1.000 0.707 gcagttaaaatTACGaatcatgg SEQ_9 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain protein AGA- 0.82 176 196 (−) 1.000 0.820 ggagcCCATgattcgtaattt MOUS-like 2 SEQ_9 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol biosynthesis 0.85 196 206 (+) 1.000 0.897 ctATCTatatt and in the expression of photosynthesis- related genes SEQ_9 P$MYBS/ZMMRP1.01 Zea mays MYB-related protein 1 (transfer cell specific) 0.79 196 212 (+) 0.777 0.905 ctatctaTATTttacat SEQ_9 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 200 216 (−) 0.761 0.834 tgtaatgtAAAAtatag protein gene promoters SEQ_9 P$DOFF/DOF2.01 Dof2 - single zinc finger transcription factor 0.98 211 227 (+) 1.000 0.988 attacaatAAAGctttt SEQ_9 P$DOFF/DOF2.01 Dof2 - single zinc finger transcription factor 0.98 242 258 (+) 1.000 0.995 attacaatAAAGctata SEQ_9 P$TBPF/TATA.02 Plant TATA box 0.90 251 265 (+) 1.000 0.913 aagcTATAtatcact SEQ_9 P$MYBS/ZMMRP1.01 Zea mays MYB-related protein 1 (transfer cell specific) 0.79 272 288 (−) 0.777 0.827 ttgtcttTATTtcagat SEQ_9 P$DOFF/DOF1.01 Dof1/MNB1a - single zinc finger transcription factor 0.98 273 289 (+) 1.000 0.984 tctgaaatAAAGacaaa SEQ_9 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 280 292 (−) 1.000 0.940 gggtTTGTcttta SEQ_9 P$TELO/RPBX.01 Ribosomal protein box, appears unique to plant RP 0.84 285 299 (+) 1.000 0.864 acaaaCCCTgaactc genes and genes associated with gene expression SEQ_9 P$AHBP/ATHB1.01 Arabidopsis thaliana homeo box protein 1 0.90 307 317 (−) 1.000 0.989 ctaATTAtttc SEQ_9 P$AHBP/HAHB4.01 Sunflower homeodomain leucine-zipper protein 0.87 307 317 (+) 1.000 0.943 gaaataATTAg Hahb-4 SEQ_9 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), AtMYC2 0.95 317 335 (+) 0.954 0.972 gccaaACACttgattccaa (rd22BP1) SEQ_9 P$MADS/SQUA.01 MADS-box protein SQUAMOSA 0.90 334 354 (−) 1.000 0.906 ggtcgctATTTgtttctgttt SEQ_9 P$ERSE/ERSE_I.01 ERSE I (ER stress-response element I)-like motif 0.79 343 361 (+) 1.000 0.799 caaatagcgacctaaCACG SEQ_9 P$IBOX/GATA.01 Class I GATA factors 0.93 356 372 (+) 1.000 0.958 aacacGATAatagatag SEQ_9 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol biosynthesis 0.85 362 372 (−) 1.000 0.853 ctATCTattat and in the expression of photosynthesis- related genes SEQ_9 P$AHBP/BLR.01 Transcriptional repressor BELLRINGER 0.90 383 393 (−) 1.000 0.928 tatATTAtttt SEQ_9 P$IBOX/IBOX.01 I-Box in rbcS genes and other light regulated genes 0.81 385 401 (+) 0.750 0.817 aataaTATAaggatcag SEQ_9 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 398 412 (−) 1.000 0.864 aaccaagaAATCtga SEQ_9 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that binds to the 0.87 411 423 (−) 1.000 0.909 ttTACTgtttaaa SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_9 P$GAPB/GAP.01 Cis-element in the GAPDH promoters conferring light 0.88 431 445 (+) 1.000 0.984 acacATGAagacaag inducibility SEQ_9 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 448 466 (+) 1.000 0.815 aaAAATtatagattttaca sugar-responsive genes SEQ_9 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol biosynthesis 0.85 452 462 (−) 1.000 0.912 aaATCTataat and in the expression of photosynthesis- related genes SEQ_9 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 455 469 (−) 1.000 0.889 ttttgtaaAATCtat SEQ_9 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 455 471 (−) 0.761 0.841 tgttttgtAAAAtctat protein gene promoters SEQ_9 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain protein AGA- 0.82 496 516 (−) 0.968 0.856 agaagCCAAcattggcaacga MOUS-like 2 SEQ_9 P$MADS/AG.01 Agamous, required for normal flower development, 0.80 497 517 (+) 0.902 0.806 cgtTGCCaatgttggcttctt similarity to SRF (human) and MCM (yeast) proteins SEQ_9 P$LFYB/LFY.01 Plant specific floral meristem identity gene LEAFY 0.93 500 512 (+) 0.885 0.938 tGCCAatgttggc (LFY) SEQ_9 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed storage 0.75 510 526 (−) 1.000 0.801 tgtggtggAAAGaagcc protein gene promoters SEQ_9 P$SALT/ALFIN1.01 Zinc-finger protein in alfalfa roots, regulates salt 0.93 516 530 (−) 1.000 0.986 tttgtGTGGtggaaa tolerance SEQ_9 P$TEFB/TEF1.01 TEF cis acting elements in both RNA polymerase II- 0.76 521 541 (−) 0.838 0.781 taACGGtcatatttgtgtggt dependent promoters and rDNA spacer sequences SEQ_9 P$WBXF/ERE.01 Elicitor response element 0.89 527 543 (+) 1.000 0.897 caaataTGACcgttaag SEQ_9 P$MYBL/ATMYB77.01 R2R3-type myb-like transcription factor (I-type binding 0.87 530 546 (+) 0.857 0.916 atatgaCCGTtaagact site) SEQ_9 P$MSAE/MSA.01 M-phase-specific activators (NtmybA1, NtmybA2, 0.80 531 545 (−) 1.000 0.889 gtcttAACGgtcata NtmybB) SEQ_9 P$AHBP/HAHB4.01 Sunflower homeodomain leucine-zipper protein 0.87 558 568 (+) 1.000 0.916 tttataATTAc Hahb-4 SEQ_9 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix DNA-binding 0.85 558 574 (−) 0.968 0.859 catgtaGTAAttataaa domain SEQ_9 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription factors that 0.81 564 580 (+) 1.000 0.823 attactACATggatgta belong to the group of Opaque-2 like proteins SEQ_9 P$HMGF/HMG_IY.01 High mobility group I/Y-like proteins 0.89 577 591 (−) 1.000 0.912 tataTATTttataca SEQ_9 P$MYBS/ZMMRP1.01 Zea mays MYB-related protein 1 (transfer cell specific) 0.79 580 596 (−) 0.777 0.793 cgatctaTATAttttat SEQ_9 P$TBPF/TATA.02 Plant TATA box 0.90 581 595 (−) 1.000 0.909 gatcTATAtatttta SEQ_9 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol biosynthesis 0.85 586 596 (−) 1.000 0.882 cgATCTatata and in the expression of photosynthesis- related genes SEQ_9 P$L1BX/PDF2.01 Protodermal factor 2 0.85 617 633 (−) 1.000 0.891 gagaaaTAAAtggtcga

TABLE 4 cis-regulatory elements of SEQ ID NO: 14 Seq. Opt. Start End Core Matrix name Family/matrix Further Information thresh. pos. pos. Strand sim. sim. Sequence SEQ_14 P$OCSE/OCSL.01 OCS-like elements 0.69 7 27 (−) 1.000 0.702 caaagtgtgactatACGTttt SEQ_14 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription 0.99 31 47 (−) 1.000 0.997 catagtcaAAAGcacaa factor SEQ_14 P$WBXF/WRKY.01 WRKY plant specific zinc-finger-type 0.92 34 50 (+) 1.000 0.961 tgcttTTGActatgtgt factor associated with pathogen defence, W box SEQ_14 P$MYBS/ZMMRP1.01 Zea mays MYB-related protein 1 (transfer 0.79 43 59 (+) 1.000 0.833 ctatgtgTATCtgttcc cell specific) SEQ_14 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 59 69 (−) 1.000 0.892 cgATCTataag biosynthesis and in the expression of photosynthesis-related genes SEQ_14 P$MADS/AG.01 Agamous, required for normal flower 0.80 92 112 (−) 0.962 0.820 tagTTCCcaaaccggttccaa development, similarity to SRF (human) and MCM (yeast) proteins SEQ_14 P$MIIG/MYBC1.01 Maize C1 myb-domain protein 0.92 104 118 (−) 1.000 0.935 tgttgGTAGttccca SEQ_14 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain 0.82 108 128 (+) 0.968 0.828 aactaCCAAcacaagcaatgc protein AGAMOUS-like 2 SEQ_14 P$NCS2/NCS2.01 Nodulin consensus sequence 2 0.79 130 144 (−) 1.000 0.817 ttttgcCTCTctaag SEQ_14 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato 0.87 139 151 (−) 1.000 0.918 atTACTcttttgc that binds to the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_14 P$DOFF/PBF.01 PBF (MPBF) 0.97 145 161 (+) 1.000 0.990 gagtaataAAAGagagg SEQ_14 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 163 173 (+) 1.000 0.862 gAAAAgttttg SEQ_14 P$WBXF/WRKY.01 WRKY plant specific zinc-finger-type 0.92 166 182 (−) 1.000 0.942 atagtTTGAcaaaactt factor associated with pathogen defence, W box SEQ_14 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger 0.92 167 179 (+) 1.000 0.935 agttTTGTcaaac protein SEQ_14 P$OCSE/OCSL.01 OCS-like elements 0.69 194 214 (−) 0.769 0.721 aaaagttagatcttACTTtct SEQ_14 P$OCSE/OCSL.01 OCS-like elements 0.69 195 215 (+) 0.769 0.766 gaaagtaagatctaACTTttt SEQ_14 P$AGP1/AGP1.01 AG-motif binding protein 1 0.91 199 209 (−) 1.000 0.915 ttaGATCttac SEQ_14 P$AGP1/AGP1.01 AG-motif binding protein 1 0.91 200 210 (+) 1.000 0.983 taaGATCtaac SEQ_14 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 202 212 (+) 1.000 0.910 agATCTaactt biosynthesis and in the expression of photosynthesis-related genes SEQ_14 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 202 218 (−) 1.000 0.819 aaaaaaaaGTTAgatct SEQ_14 P$MYBL/NTMYBAS1.01 Anther-specific myb gene from tobacco 0.96 222 238 (+) 1.000 0.976 tttgggatGTTAggctt SEQ_14 P$TELO/ATPURA.01 Arabidopsis Telo-box interacting protein 0.85 227 241 (−) 0.750 0.868 caaaAGCCtaacatc related to the conserved animal protein Pur-alpha SEQ_14 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription 0.99 231 247 (−) 1.000 0.997 agccttcaAAAGcctaa factor SEQ_14 P$SUCB/SUCROSE.01 Sequence motif from the promoters of 0.81 238 256 (−) 0.750 0.836 aaAAAAcatagccttcaaa different sugar-responsive genes SEQ_14 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 268 294 (−) 0.750 0.593 accttcgACATgatctaagaacaaaga element about 100 bp up- stream of the TSS in legumin genes SEQ_14 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 274 284 (−) 1.000 0.921 tgATCTaagaa biosynthesis and in the expression of photosynthesis-related genes SEQ_14 P$CARM/CARICH.01 CA-rich element 0.78 286 304 (−) 1.000 0.816 acatttcAACAccttcgac SEQ_14 P$L1BX/PDF2.01 Protodermal factor 2 0.85 308 324 (+) 1.000 0.903 atgtaaTAAAtgttatt SEQ_14 P$OCSE/OCSL.01 OCS-like elements 0.69 314 334 (−) 0.807 0.714 gcagctgagtaataACATtta SEQ_14 P$OCSE/OCSL.01 OCS-like elements 0.69 322 342 (+) 1.000 0.717 attactcagctgctACGTtta SEQ_14 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 368 382 (−) 1.000 0.852 tttacataAATCtca SEQ_14 P$DOFF/PBOX.01 Prolamin box, conserved in cereal seed 0.75 372 388 (+) 0.761 0.751 atttatgtAAAAtccat storage protein gene promoters SEQ_14 P$IBOX/GATA.01 Class I GATA factors 0.93 399 415 (−) 1.000 0.973 aaatgGATAagattgat SEQ_14 P$MYBS/HVMCB1.01 Hordeum vulgare Myb-related CAB- 0.93 402 418 (+) 1.000 0.963 aatcttATCCattttct promoter-binding protein 1 SEQ_14 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 420 430 (+) 1.000 0.963 ctgatTAATct SEQ_14 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 421 431 (−) 1.000 0.963 cagatTAATca SEQ_14 P$ABRE/ABRE.01 ABA response elements 0.82 428 444 (−) 1.000 0.850 taattgcACGTtgcaga SEQ_14 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 437 453 (+) 1.000 0.782 tgcaatTAGTttgatca SEQ_14 P$TEFB/TEF1.01 TEF cis acting elements in both RNA 0.76 450 470 (−) 0.838 0.779 ccATGGctaatattgtttgat polymerase II-dependent promoters and rDNA spacer sequences SEQ_14 P$GBOX/UPRE.01 UPRE (unfolded protein response 0.86 493 513 (−) 0.767 0.862 cttcgtCCAAgtcaacataag element) like motif SEQ_14 P$RAV5/RAV1-5.01 5′-part of bipartite RAV1 binding site, 0.96 493 503 (−) 1.000 0.960 gtcAACAtaag interacting with AP2 domain SEQ_14 P$GBOX/BZIP911.01 bZIP transcription factor from Antirrhinum 0.77 494 514 (+) 1.000 0.833 ttatgtTGACttggacgaaga majus SEQ_14 P$OPAQ/O2.01 Opaque-2 regulatory protein 0.87 495 511 (−) 0.852 0.895 tcgtccaagTCAAcata SEQ_14 P$DOFF/DOF2.01 Dof2 - single zinc finger transcription 0.98 544 560 (−) 1.000 1.000 tatttattAAAGcaaac factor SEQ_14 P$L1BX/ATML1.01 L1-specific homeodomain protein 0.82 549 565 (+) 1.000 0.853 ctttaaTAAAtataagt ATML1 (A. thaliana meristem layer 1) SEQ_14 P$SUCB/SUCROSE.01 Sequence motif from the promoters of 0.81 559 577 (−) 0.750 0.816 caCAATcattctacttata different sugar-responsive genes SEQ_14 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 566 576 (+) 0.829 0.904 agaATGAttgt SEQ_14 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 566 576 (−) 0.936 0.977 acaATCAttct SEQ_14 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger 0.92 575 587 (+) 1.000 0.926 gtgtTTGTcttct protein SEQ_14 P$GTBX/SBF1.01 SBF-1 0.87 581 597 (−) 1.000 0.885 tctgtgaTTAAgaagac SEQ_14 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 583 593 (+) 1.000 0.963 cttctTAATca SEQ_14 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato 0.87 590 602 (−) 1.000 0.876 aaTACTctgtgat that binds to the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_14 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 610 620 (−) 1.000 0.963 aacctTAATct SEQ_14 P$OCSE/OCSL.01 OCS-like elements 0.69 614 634 (+) 1.000 0.699 taaggtttgaatgaACGTcgt SEQ_14 P$EINL/TEIL.01 TEIL (tobacco EIN3-like) 0.92 624 632 (+) 0.964 0.932 aTGAAcgtc SEQ_14 P$GBOX/TGA1.01 Arabidopsis leucine zipper protein 0.90 633 653 (+) 1.000 0.903 gtaaaaTGACggttatgctcg TGA1 SEQ_14 P$MYBL/ATMYB77.01 R2R3-type myb-like transcription factor 0.87 636 652 (+) 1.000 0.970 aaatgaCGGTtatgctc (I-type binding site) SEQ_14 P$TEFB/TEF1.01 TEF cis acting elements in both RNA 0.76 639 659 (+) 0.838 0.778 tgACGGttatgctcgtgagag polymerase II-dependent promoters and rDNA spacer sequences SEQ_14 P$PREM/MGPROTORE.01 Promoter elements involved in MgProto 0.77 641 671 (−) 1.000 0.792 taagCGACgattctctcacgagcataaccgt (Mg-protoporphyrin IX) and light- mediated induction SEQ_14 P$IBOX/GATA.01 Class I GATA factors 0.93 668 684 (+) 1.000 0.982 cttacGATAaggacgaa SEQ_14 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 690 706 (+) 1.000 0.936 atttgattGTTAtcagg SEQ_14 P$MYBL/NTMYBAS1.01 Anther-specific myb gene from tobacco 0.96 701 717 (+) 1.000 0.967 atcaggttGTTAaaagt SEQ_14 P$GTBX/SBF1.01 SBF-1 0.87 703 719 (+) 1.000 0.921 caggttgTTAAaagttg SEQ_14 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 711 721 (+) 1.000 0.861 tAAAAgttgag SEQ_14 P$AREF/ARE.01 Auxin Response Element 0.93 715 727 (−) 1.000 0.932 gttTGTCtcaact SEQ_14 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger 0.92 717 729 (−) 1.000 0.927 tcgtTTGTctcaa protein SEQ_14 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), 0.95 747 765 (−) 0.954 0.972 acgtaACACctgtttagtc AtMYC2 (rd22BP1) SEQ_14 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), 0.95 748 766 (+) 0.863 0.954 actaaACAGgtgttacgtt AtMYC2 (rd22BP1) SEQ_14 P$GBOX/HBP1B.01 Wheat bZIP transcription factor HBP1B 0.83 753 773 (−) 1.000 0.857 aatgtgaaACGTaacacctgt (histone gene binding protein 1b) SEQ_14 P$GTBX/GT3A.01 Trihelix DNA-binding factor GT-3a 0.83 753 769 (+) 1.000 0.858 acaggtGTTAcgtttca SEQ_14 P$ABRE/ABRE.01 ABA response elements 0.82 755 771 (+) 1.000 0.820 aggtgttACGTttcaca SEQ_14 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 755 777 (−) 0.812 0.708 aaccaatgtgaAACGtaacacct SEQ_14 P$LFYB/LFY.01 Plant specific floral meristem identity 0.93 765 777 (−) 0.914 0.947 aACCAatgtgaaa gene LEAFY (LFY) SEQ_14 P$CAAT/CAAT.01 CCAAT-box in plant promoters 0.97 769 777 (−) 1.000 0.982 aaCCAAtgt SEQ_14 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix 0.85 790 806 (+) 1.000 0.906 cttgaaGTTActctatt DNA-binding domain SEQ_14 P$AHBP/BLR.01 Transcriptional repressor BELL- 0.90 793 803 (+) 0.826 0.914 gaaGTTActct RINGER SEQ_14 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 802 818 (−) 1.000 0.828 ttggaccgGTTAaatag SEQ_14 P$WBXF/ERE.01 Elicitor response element 0.89 824 840 (−) 1.000 0.894 ttaaccTGACcggttgg SEQ_14 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix 0.85 830 846 (+) 1.000 0.854 ggtcagGTTAacaaaac DNA-binding domain SEQ_14 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 833 849 (−) 1.000 0.930 agtgttttGTTAacctg SEQ_14 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 845 861 (+) 1.000 0.910 acactgaaGTTAgccgc SEQ_14 P$GCCF/GCC-BOX.01 GCC-box, ethylene-responsive element 0.86 853 865 (+) 1.000 1.000 gttAGCCgccaac (ERE) SEQ_14 P$GBOX/HBP1B.01 Wheat bZIP transcription factor HBP1B 0.83 867 887 (+) 1.000 0.842 cgcttattACGTaaacggtag (histone gene binding protein 1b) SEQ_14 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 868 890 (−) 1.000 0.728 tggctaccgttTACGtaataagc SEQ_14 P$OCSE/OCSL.01 OCS-like elements 0.69 872 892 (−) 1.000 0.710 cgtggctaccgtttACGTaat SEQ_14 P$MSAE/MSA.01 M-phase-specific activators (NtmybA1, 0.80 875 889 (+) 1.000 0.827 acgtaAACGgtagcc NtmybA2, NtmybB) SEQ_14 P$GBOX/GBF1.01 bZIP protein G-Box binding factor 1 0.94 881 901 (−) 1.000 0.943 tgctcgaaACGTggctaccgt SEQ_14 P$GBOX/HBP1A.01 HBP-1a, suggested to be involved in 0.88 882 902 (+) 1.000 0.943 cggtagcCACGtttcgagcac the cell cycle-dependent expression SEQ_14 P$MYCL/MYCRS.01 Myc recognition sequences 0.93 882 900 (−) 1.000 0.936 gctcgaaACGTggctaccg SEQ_14 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 883 905 (−) 0.812 0.717 gcagtgctcgaAACGtggctacc SEQ_14 P$ABRE/ABRE.01 ABA response elements 0.82 884 900 (−) 1.000 0.864 gctcgaaACGTggctac SEQ_14 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger 0.92 912 924 (+) 1.000 0.921 taatTTGTcttca protein SEQ_14 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 975 1001 (−) 0.750 0.657 tcactagCCTTgcatgcgaatcagtag element about 100 bp up- stream of the TSS in legumin genes SEQ_14 P$LEGB/RY.01 RY and Sph motifs conserved in seed- 0.87 978 1004 (+) 1.000 0.899 ctgattcgCATGcaaggctagtgacac specific promoters

TABLE 5 cis-regulatory elements of SEQ ID NO: 16 Seq. Opt. Start End Core Matrix name Family/matrix Further Information thresh. pos. pos. Strand sim. sim. Sequence SEQ_16 P$OCSE/OCSL.01 OCS-like elements 0.69 7 27 (−) 1.000 0.702 caaagtgtgactatACGTttt SEQ_16 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription factor 0.99 31 47 (−) 1.000 0.997 catagtcaAAAGcacaa SEQ_16 P$WBXF/WRKY.01 WRKY plant specific zinc-finger-type factor 0.92 34 50 (+) 1.000 0.961 tgcttTTGActatgtgt associated with pathogen defence, W box SEQ_16 P$MYBS/ZMMRP1.01 Zea mays MYB-related protein 1 (transfer cell 0.79 43 59 (+) 1.000 0.833 ctatgtgTATCtgttcc specific) SEQ_16 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 59 69 (−) 1.000 0.892 cgATCTataag biosynthesis and in the expression of photo- synthesis-related genes SEQ_16 P$MADS/AG.01 Agamous, required for normal flower development, 0.80 92 112 (−) 0.962 0.820 tagTTCCcaaaccggttccaa similarity to SRF (human) and MCM (yeast) proteins SEQ_16 P$MIIG/MYBC1.01 Maize C1 myb-domain protein 0.92 104 118 (−) 1.000 0.935 tgttgGTAGttccca SEQ_16 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain protein 0.82 108 128 (+) 0.968 0.828 aactaCCAAcacaagcaatgc AGAMOUS-like 2 SEQ_16 P$GAGA/BPC.01 Basic pentacysteine proteins 1.00 129 153 (+) 1.000 1.000 tcttagAGAGagaaaagagtaataa SEQ_16 P$GAGA/BPC.01 Basic pentacysteine proteins 1.00 131 155 (+) 1.000 1.000 ttagagAGAGaaaagagtaataaaa SEQ_16 P$DOFF/PBF.01 PBF (MPBF) 0.97 134 150 (+) 1.000 0.988 gagagagaAAAGagtaa SEQ_16 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that 0.87 139 151 (−) 1.000 0.919 atTACTcttttct binds to the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta- amylase genes SEQ_16 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 140 150 (+) 1.000 0.855 gAAAAgagtaa SEQ_16 P$DOFF/PBF.01 PBF (MPBF) 0.97 145 161 (+) 1.000 0.990 gagtaataAAAGagagg SEQ_16 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 163 173 (+) 1.000 0.862 gAAAAgttttg SEQ_16 P$WBXF/WRKY.01 WRKY plant specific zinc-finger-type factor 0.92 166 182 (−) 1.000 0.942 agagtTTGAcaaaactt associated with pathogen defence, W box SEQ_16 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 167 179 (+) 1.000 0.935 agttTTGTcaaac SEQ_16 P$OCSE/OCSL.01 OCS-like elements 0.69 194 214 (−) 0.769 0.708 gaaagttagatcttACTTtct SEQ_16 P$OCSE/OCSL.01 OCS-like elements 0.69 195 215 (+) 0.769 0.733 gaaagtaagatctaACTTtct SEQ_16 P$AGP1/AGP1.01 AG-motif binding protein 1 0.91 199 209 (−) 1.000 0.915 ttaGATCttac SEQ_16 P$AGP1/AGP1.01 AG-motif binding protein 1 0.91 200 210 (+) 1.000 0.983 taaGATCtaac SEQ_16 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 202 212 (+) 1.000 0.910 agATCTaactt biosynthesis and in the expression of photo- synthesis-related genes SEQ_16 P$MYBL/NTMYBAS1.01 Anther-specific myb gene from tobacco 0.96 228 244 (+) 1.000 0.976 tttgggatGTTAggctt SEQ_16 P$TELO/ATPURA.01 Arabidopsis Telo-box interacting protein re- 0.85 233 247 (−) 0.750 0.868 caaaAGCCtaacatc lated to the conserved animal protein Pur- alpha SEQ_16 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription factor 0.99 237 253 (−) 1.000 0.997 agccttcaAAAGcctaa SEQ_16 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 244 262 (−) 0.750 0.836 aaAAAAcatagccttcaaa sugar-responsive genes SEQ_16 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 274 300 (−) 0.750 0.593 accttcgACATgatctaagaacaaaga element about 100 bp upstream of the TSS in legumin genes SEQ_16 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol 0.85 280 290 (−) 1.000 0.921 tgATCTaagaa biosynthesis and in the expression of photo- synthesis-related genes SEQ_16 P$CARM/CARICH.01 CA-rich element 0.78 292 310 (−) 1.000 0.787 acacttcAACAccttcgac SEQ_16 P$CARM/CARICH.01 CA-rich element 0.78 300 318 (−) 1.000 0.829 tacgtccAACActtcaaca SEQ_16 P$OCSE/OCSL.01 OCS-like elements 0.69 311 331 (−) 1.000 0.695 taataacatttattACGTcca SEQ_16 P$L1BX/PDF2.01 Protodermal factor 2 0.85 314 330 (+) 1.000 0.903 acgtaaTAAAtgttatt SEQ_16 P$OCSE/OCSL.01 OCS-like elements 0.69 320 340 (−) 0.807 0.714 gcagctgagtaataACATtta SEQ_16 P$OCSE/OCSL.01 OCS-like elements 0.69 328 348 (+) 1.000 0.717 attactcagctgctACGTtta SEQ_16 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 374 388 (−) 1.000 0.852 tttacataAATCtca SEQ_16 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 386 404 (+) 1.000 0.818 aaAAATccgcaatcttatc sugar-responsive genes SEQ_16 P$IBOX/GATA.01 Class I GATA factors 0.93 393 409 (−) 1.000 0.973 aaatgGATAagattgcg SEQ_16 P$MYBS/HVMCB1.01 Hordeum vulgare Myb-related CAB-promoter- 0.93 396 412 (+) 1.000 0.963 aatcttATCCattttct binding protein 1 SEQ_16 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 414 424 (+) 1.000 0.963 ctgatTAATct SEQ_16 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 415 425 (−) 1.000 0.963 cagatTAATca SEQ_16 P$ABRE/ABRE.01 ABA response elements 0.82 422 438 (−) 1.000 0.850 taattgcACGTtgcaga SEQ_16 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 431 447 (+) 1.000 0.782 tgcaatTAGTttgatca SEQ_16 P$HMGF/HMG_IY.01 High mobility group I/Y-like proteins 0.89 441 455 (−) 1.000 0.907 atatTATTtgatcaa SEQ_16 P$AHBP/BLR.01 Transcriptional repressor BELLRINGER 0.90 446 456 (−) 1.000 0.928 aatATTAtttg SEQ_16 P$RAV5/RAV1-5.01 5′-part of bipartite RAV1 binding site, interacting 0.96 487 497 (−) 1.000 0.960 gtcAACAtaag with AP2 domain SEQ_16 P$OPAQ/O2.01 Opaque-2 regulatory protein 0.87 489 505 (−) 0.852 0.895 tcctccaagTCAAcata SEQ_16 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription factor 0.99 538 554 (−) 1.000 0.994 tatttataAAAGcaaac SEQ_16 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 538 554 (−) 1.000 0.859 tatttaTAAAagcaaac meristem layer 1) SEQ_16 P$L1BX/ATML1.01 L1-specific homeodomain protein ATML1 (A. thaliana 0.82 543 559 (+) 1.000 0.834 cttttaTAAAtataagt meristem layer 1) SEQ_16 P$TBPF/TATA.01 Plant TATA box 0.88 543 557 (+) 1.000 0.971 ctttTATAaatataa SEQ_16 P$SUCB/SUCROSE.01 Sequence motif from the promoters of different 0.81 553 571 (−) 0.750 0.816 caCAATcattctacttata sugar-responsive genes SEQ_16 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 560 570 (+) 0.829 0.904 agaATGAttgt SEQ_16 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 560 570 (−) 0.936 0.977 acaATCAttct SEQ_16 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 569 581 (+) 1.000 0.926 gtgtTTGTcttct SEQ_16 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato that 0.87 586 598 (−) 1.000 0.876 aaTACTctgtgat binds to the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta- amylase genes SEQ_16 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 606 616 (−) 1.000 0.963 aacctTAATct SEQ_16 P$GBOX/TGA1.01 Arabidopsis leucine zipper protein TGA1 0.90 625 645 (+) 1.000 0.903 gtaaaaTGACggttatgctcg SEQ_16 P$MYBL/ATMYB77.01 R2R3-type myb-like transcription factor (I-type 0.87 628 644 (+) 1.000 0.970 aaatgaCGGTtatgctc binding site) SEQ_16 P$TEFB/TEF1.01 TEF cis acting elements in both RNA polymerase 0.76 631 651 (+) 0.838 0.778 tgACGGttatgctcgtgagag II-dependent promoters and rDNA spacer sequences SEQ_16 P$IBOX/GATA.01 Class I GATA factors 0.93 660 676 (+) 1.000 0.950 cttgcGATAaggacgaa SEQ_16 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 682 698 (+) 1.000 0.986 atttggttGTTAtcagg SEQ_16 P$MIIG/PALBOXL.01 Cis-acting element conserved in various PAL 0.80 693 707 (+) 0.750 0.818 atcaggttGTTGaaa and 4CL promoters SEQ_16 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 693 709 (+) 0.817 0.777 atcaggTTGTtgaaaga SEQ_16 P$NCS2/NCS2.01 Nodulin consensus sequence 2 0.79 705 719 (−) 0.750 0.801 gtttgtCTCAtcttt GCN4, conserved in cereal seed storage protein gene promoters, similar to yeast SEQ_16 P$OPAQ/GCN4.01 GCN4 and vertebrate AP-1 0.81 705 721 (+) 1.000 0.846 aaagaTGAGacaaacga SEQ_16 P$AREF/ARE.01 Auxin Response Element 0.93 707 719 (−) 1.000 0.951 gttTGTCtcatct SEQ_16 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 709 721 (−) 1.000 0.927 tcgtTTGTctcat SEQ_16 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), AtMYC2 0.95 739 757 (−) 0.954 0.972 acgtaACACctgtttagtc (rd22BP1) SEQ_16 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), AtMYC2 0.95 740 758 (+) 0.863 0.954 actaaACAGgtgttacgtt (rd22BP1) SEQ_16 P$GBOX/HBP1B.01 Wheat bZIP transcription factor HBP1B (histone 0.83 745 765 (−) 1.000 0.857 aatgtgaaACGTaacacctgt gene binding protein 1b) SEQ_16 P$GTBX/GT3A.01 Trihelix DNA-binding factor GT-3a 0.83 745 761 (+) 1.000 0.858 acaggtGTTAcgtttca SEQ_16 P$ABRE/ABRE.01 ABA response elements 0.82 747 763 (+) 1.000 0.820 aggtgttACGTttcaca SEQ_16 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 747 769 (−) 0.812 0.708 aaccaatgtgaAACGtaacacct SEQ_16 P$LFYB/LFY.01 Plant specific floral meristem identity gene 0.93 757 769 (−) 0.914 0.947 aACCAatgtgaaa LEAFY (LFY) SEQ_16 P$CAAT/CAAT.01 CCAAT-box in plant promoters 0.97 761 769 (−) 1.000 0.982 aaCCAAtgt SEQ_16 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 794 810 (−) 1.000 0.818 ccggaccgGTTAaatag SEQ_16 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 809 821 (−) 1.000 0.929 agttTTGTcaacc SEQ_16 P$GCCF/ERE_JERE.01 Ethylene-responsive elements (ERE) and 0.85 826 838 (+) 1.000 0.907 aaaaggCGCCaac jasmonate- and elicitor-responsive elements (JERE) SEQ_16 P$GBOX/HBP1B.01 Wheat bZIP transcription factor HBP1B (histone 0.83 840 860 (+) 1.000 0.858 cgcttgttACGTaaacggtag gene binding protein 1b) SEQ_16 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 841 863 (−) 1.000 0.764 tggctaccgttTACGtaacaagc SEQ_16 P$OCSE/OCSL.01 OCS-like elements 0.69 845 865 (−) 1.000 0.710 cgtggctaccgtttACGTaac SEQ_16 P$MSAE/MSA.01 M-phase-specific activators (NtmybA1, 0.80 848 862 (+) 1.000 0.827 acgtaAACGgtagcc NtmybA2, NtmybB) SEQ_16 P$GBOX/GBF1.01 bZIP protein G-Box binding factor 1 0.94 854 874 (−) 1.000 0.944 tgctcaaaACGTggctaccgt SEQ_16 P$GBOX/HBP1A.01 HBP-1a, suggested to be involved in the cell 0.88 855 875 (+) 1.000 0.914 cggtagcCACGttttgagcac cycle-dependent expression SEQ_16 P$NACF/TANAC69.01 Wheat NACdomain DNA binding factor 0.68 856 878 (−) 0.812 0.712 gcagtgctcaaAACGtggctacc SEQ_16 P$ABRE/ABRE.01 ABA response elements 0.82 857 873 (−) 1.000 0.860 gctcaaaACGTggctac SEQ_16 P$MYCL/MYCRS.01 Myc recognition sequences 0.93 872 890 (+) 0.863 0.946 gcactgcATGTgctaattt SEQ_16 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger protein 0.92 885 897 (+) 1.000 0.921 taatTTGTcttca SEQ_16 P$TCPF/ATTCP20.01 TCP class I transcription factor (Arabidopsis) 0.94 924 936 (−) 1.000 0.947 ttaaGCCCaagtg SEQ_16 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 940 956 (−) 1.000 0.781 attaatTAGTtccacga SEQ_16 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 948 958 (+) 1.000 1.000 ctaatTAATga SEQ_16 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 952 962 (+) 0.829 0.902 ttaATGAttcg SEQ_16 P$AHBP/HAHB4.01 Sunflower homeodomain leucine-zipper protein 0.87 952 962 (−) 1.000 0.979 cgaatcATTAa Hahb-4 SEQ_16 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 952 978 (−) 0.750 0.657 tcactagCCTTgcatgcgaatcattaa element about 100 by upstream of the TSS in legumin genes SEQ_16 P$LEGB/RY.01 RY and Sph motifs conserved in seed-specific 0.87 955 981 (+) 1.000 0.899 atgattcgCATGcaaggctagtgacac promoters

TABLE 6 cis-regulatory elements of SEQ ID NO: 22 Seq. Opt. Start End Core Matrix name Family/matrix Further Information thresh. pos. pos. Strand sim. sim. Sequence SEQ_22 P$AHBP/BLR.01 Transcriptional repressor BELL- 0.90 4 14 (+) 0.826 0.936 gaaGTTAttag RINGER SEQ_22 P$MYBL/CARE.01 CAACTC regulatory elements, GA- 0.83 27 43 (−) 1.000 0.875 gttggctAGTTgtaagt inducible SEQ_22 P$WBXF/WRKY.01 WRKY plant specific zinc-finger-type 0.92 49 65 (−) 1.000 0.975 catgtTTGAcctctaca factor associated with pathogen defence, W box SEQ_22 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription 0.81 56 72 (−) 1.000 0.853 tttgtaACATgtttgac factors that belong to the group of Opaque-2 like proteins SEQ_22 P$GTBX/GT3A.01 Trihelix DNA-binding factor GT-3a 0.83 59 75 (+) 1.000 0.867 aaacatGTTAcaaactc SEQ_22 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 61 77 (−) 0.817 0.798 ttgagtTTGTaacatgt SEQ_22 P$HMGF/HMG_IY.02 High mobility group I/Y-like protein 1.00 72 86 (−) 1.000 1.000 ctttTATTtttgagt isolated from pea SEQ_22 P$PSRE/GAAA.01 GAAA motif involved in pollen specific 0.83 81 97 (+) 1.000 0.879 taaaaGAAAcagtggag transcriptional activation SEQ_22 P$LFYB/LFY.01 Plant specific floral meristem identity 0.93 85 97 (−) 1.000 0.969 cTCCActgtttct gene LEAFY (LFY) SEQ_22 P$GTBX/GT1.01 GT1-Box binding factors with a trihelix 0.85 100 116 (−) 1.000 0.904 gttcagGTTActcgatt DNA-binding domain SEQ_22 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), 0.95 109 127 (−) 0.954 0.972 atcaaACACctgttcaggt AtMYC2 (rd22BP1) SEQ_22 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), 0.95 110 128 (+) 0.863 0.954 cctgaACAGgtgtttgatc AtMYC2 (rd22BP1) SEQ_22 P$CARM/CARICH.01 CA-rich element 0.78 112 130 (−) 1.000 0.809 ttgatcaAACAcctgttca SEQ_22 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 140 156 (−) 1.000 0.781 aagagaTAGTgacacac SEQ_22 P$MYBS/ZMMRP1.01 Zea mays MYB-related protein 1 0.79 142 158 (+) 1.000 0.824 gtgtcacTATCtcttgg (transfer cell specific) SEQ_22 P$DOFF/DOF1.01 Dof1/MNB1a - single zinc finger 0.98 169 185 (+) 1.000 0.984 acacaaatAAAGaccct transcription factor SEQ_22 P$MIIG/PALBOXL.01 Cis-acting element conserved in 0.80 188 202 (−) 0.785 0.801 agcagcttGGTTagg various PAL and 4CL promoters SEQ_22 P$CAAT/CAAT.01 CCAAT-box in plant promoters 0.97 204 212 (+) 1.000 0.985 atCCAAtcc SEQ_22 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription 0.81 206 222 (−) 0.829 0.819 tgtgtgACTTggattgg factors that belong to the group of Opaque-2 like proteins SEQ_22 P$L1BX/PDF2.01 Protodermal factor 2 0.85 229 245 (+) 1.000 0.921 cagctaTAAAtgaaaca SEQ_22 P$TBPF/TATA.02 Plant TATA box 0.90 229 243 (+) 1.000 0.935 cagcTATAaatgaaa SEQ_22 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 250 266 (−) 1.000 0.808 attcatctGTTAaagtt SEQ_22 P$GAPB/GAP.01 Cis-element in the GAPDH promoters 0.88 257 271 (+) 1.000 0.964 acagATGAatactag conferring light inducibility SEQ_22 P$HEAT/HSE.01 Heat shock element 0.81 284 298 (−) 1.000 0.811 aggagacactAGAAc SEQ_22 P$AREF/ARE.01 Auxin Response Element 0.93 288 300 (+) 1.000 0.961 tagTGTCtcctca SEQ_22 P$NCS2/NCS2.01 Nodulin consensus sequence 2 0.79 288 302 (+) 0.750 0.819 tagtgtCTCCtcatt SEQ_22 P$ROOT/RHE.01 Root hair-specific element with a 2- 0.77 298 322 (−) 1.000 0.852 atcgtagcttgaattCACGtaatga nucleotid spacer between left part (LP) and right part (RP) SEQ_22 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 317 333 (−) 1.000 0.775 ttgagaTAGTgatcgta SEQ_22 P$MYBL/CARE.01 CAACTC regulatory elements, GA- 0.83 326 342 (−) 1.000 0.838 atgtaggAGTTgagata inducible SEQ_22 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 357 383 (+) 0.750 0.595 tacaaaaCTATgcacaaaaacaaaagc element about 100 by up- stream of the TSS in legumin genes SEQ_22 P$EINL/TEIL.01 TEIL (tobacco EIN3-like) 0.92 380 388 (−) 1.000 0.934 aTGTAgctt SEQ_22 P$LREM/ATCTA.01 Motif involved in carotenoid and tocopherol biosynthesis and in the expression of photosynthesis-related 0.85 385 395 (+) 1.000 0.931 acATCTaatac genes SEQ_22 P$CE1F/ABI4.01 ABA insensitive protein 4 (ABI4) 0.87 432 444 (+) 1.000 0.872 caatCACCgtcga SEQ_22 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 450 466 (+) 1.000 0.849 aggattcaGTTAattga SEQ_22 P$CGCG/OSCBT.01 Oryza sativa CaM-binding transcription 0.78 475 491 (−) 0.817 0.796 cttCGAGtttgatcgga factor SEQ_22 P$ROOT/RHE.02 Root hair-specific element with a 3- 0.77 486 510 (+) 1.000 0.801 tcgaagactggtgagCACGaggacg nucleotid spacer between left part (LP) and right part (RP) SEQ_22 P$NCS2/NCS2.01 Nodulin consensus sequence 2 0.79 525 539 (−) 0.750 0.823 tgttgtATCTtcgag SEQ_22 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 539 553 (+) 1.000 0.859 aagcaagaAATCtac SEQ_22 P$LREM/ATCTA.01 Motif involved in carotenoid and 0.85 546 556 (+) 1.000 0.869 aaATCTactga tocopherol biosynthesis and in the expression of photosynthesis-related genes SEQ_22 P$L1BX/ATML1.02 Arabidopsis thaliana meristem layer 1 0.76 563 579 (−) 0.890 0.767 cgcCAATaacttcagga SEQ_22 P$AHBP/BLR.01 Transcriptional repressor BELL- 0.90 567 577 (+) 0.826 0.936 gaaGTTAttgg RINGER SEQ_22 P$L1BX/HDG9.01 Homeodomain glabrous 9 0.77 595 611 (−) 0.796 0.777 ccgaaaTTAAttcggat SEQ_22 P$L1BX/HDG9.01 Homeodomain glabrous 9 0.77 597 613 (+) 0.750 0.796 ccgaatTAATttcgggg SEQ_22 P$AHBP/BLR.01 Transcriptional repressor BELL- 0.90 599 609 (−) 1.000 1.000 gaaATTAattc RINGER SEQ_22 P$IBOX/GATA.01 Class I GATA factors 0.93 614 630 (+) 1.000 0.961 aaaaaGATAaattagat SEQ_22 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 614 624 (+) 1.000 0.948 aAAAAgataaa SEQ_22 P$LREM/ATCTA.01 Motif involved in carotenoid and 0.85 622 632 (−) 1.000 0.921 gtATCTaattt tocopherol biosynthesis and in the expression of photosynthesis-related genes SEQ_22 P$ABRE/ABF1.01 ABA (abscisic acid) inducible transcriptional 0.79 657 673 (+) 0.750 0.797 aagaaACAGgtggcaat activator SEQ_22 P$TCPF/ATTCP20.01 TCP class I transcription factor 0.94 670 682 (−) 1.000 0.949 tccaGCCCaattg (Arabidopsis) SEQ_22 P$OCSE/OCSL.01 OCS-like elements 0.69 715 735 (+) 0.807 0.693 aaaaaaaacggataACATatt SEQ_22 P$MSAE/MSA.01 M-phase-specific activators 0.80 716 730 (+) 1.000 0.851 aaaaaAACGgataac (NtmybA1, NtmybA2, NtmybB) SEQ_22 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 717 733 (+) 0.750 0.845 aaaaaacgGATAacata SEQ_22 P$MYBS/MYBST1.01 MybSt1 (Myb Solanum tuberosum 1) 0.90 717 733 (−) 1.000 0.962 tatgttATCCgtttttt with a single myb repeat SEQ_22 P$IBOX/GATA.01 Class I GATA factors 0.93 720 736 (+) 1.000 0.935 aaacgGATAacatattt SEQ_22 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 722 738 (−) 1.000 0.920 ataaatatGTTAtccgt SEQ_22 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 764 778 (−) 0.757 0.857 aaaaagaaAATAtct SEQ_22 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 771 787 (+) 1.000 0.914 ttctttttGTTAggaaa SEQ_22 P$SEF4/SEF4.01 Soybean embryo factor 4 0.98 772 782 (+) 1.000 0.981 tcTTTTtgtta SEQ_22 P$HEAT/HSE.01 Heat shock element 0.81 783 797 (+) 1.000 0.873 ggaaaattttAGAAa SEQ_22 P$HMGF/HMG_IY.01 High mobility group I/Y-like proteins 0.89 790 804 (−) 1.000 0.895 ccatTATTttctaaa SEQ_22 P$SUCB/SUCROSE.01 Sequence motif from the promoters 0.81 794 812 (+) 1.000 0.851 gaAAATaatggaaattaaa of different sugar-responsive genes SEQ_22 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 795 805 (−) 1.000 0.918 tccATTAtttt SEQ_22 P$GTBX/S1F.01 S1F, site 1 binding factor of spinach 0.79 797 813 (+) 1.000 0.841 aataATGGaaattaaat rps1 promoter SEQ_22 P$GTBX/SBF1.01 SBF-1 0.87 801 817 (+) 1.000 0.908 atggaaaTTAAatagcg SEQ_22 P$L1BX/HDG9.01 Homeodomain glabrous 9 0.77 803 819 (+) 1.000 0.822 ggaaatTAAAtagcgat SEQ_22 P$GTBX/GT3A.01 Trihelix DNA-binding factor GT-3a 0.83 817 833 (+) 1.000 0.843 gattatGTTAcaagata SEQ_22 P$OCSE/OCSL.01 OCS-like elements 0.69 819 839 (+) 0.807 0.707 ttatgttacaagatACGAtca SEQ_22 P$GARP/ARR10.01 Type-B response regulator (ARR10), 0.97 829 837 (+) 1.000 0.985 AGATacgat member of the GARP-family of plant myb-related DNA binding motifs SEQ_22 P$ROOT/RHE.02 Root hair-specific element with a 3- 0.77 838 862 (−) 0.750 0.777 tagcattttgcactgCCCGatgctg nucleotid spacer between left part (LP) and right part (RP) SEQ_22 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 877 887 (−) 0.804 0.899 aAAAGgatcaa SEQ_22 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 882 898 (+) 1.000 0.926 ccttttggGTTAtctcc SEQ_22 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 903 919 (+) 1.000 0.785 gacaatTAGTttaggat SEQ_22 P$STKM/STK.01 Storekeeper (STK), plant specific 0.85 904 918 (−) 1.000 0.865 tccTAAActaattgt DNA binding protein important for tuber-specific and sucrose-inducible gene expression SEQ_22 P$TELO/ATPURA.01 Arabidopsis Telo-box interacting 0.85 909 923 (−) 0.750 0.867 caaaATCCtaaacta protein related to the conserved animal protein Pur-alpha SEQ_22 P$AHBP/BLR.01 Transcriptional repressor BELL- 0.90 929 939 (+) 1.000 0.930 tatATTAatac RINGER SEQ_22 P$IBOX/GATA.01 Class I GATA factors 0.93 935 951 (−) 1.000 0.932 tgtcgGATAatagtatt SEQ_22 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato 0.87 935 947 (+) 1.000 0.995 aaTACTattatcc that binds to the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_22 P$MYBS/MYBST1.01 MybSt1 (Myb Solanum tuberosum 1) 0.90 938 954 (+) 1.000 0.943 actattATCCgacaaca with a single myb repeat SEQ_22 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger 0.92 944 956 (−) 1.000 0.922 agtgTTGTcggat protein SEQ_22 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), 0.95 946 964 (−) 0.954 0.972 ctgaaACAAgtgttgtcgg AtMYC2 (rd22BP1) SEQ_22 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), 0.95 947 965 (+) 0.954 0.984 cgacaACACttgtttcagc AtMYC2 (rd22BP1) SEQ_22 P$GTBX/GT3A.01 Trihelix DNA-binding factor GT-3a 0.83 968 984 (−) 1.000 0.839 aaaaatGTTAaaataag SEQ_22 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 970 986 (−) 1.000 0.807 caaaaaatGTTAaaata SEQ_22 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 975 989 (−) 0.766 0.872 aaacaaaaAATGtta SEQ_22 P$GTBX/S1F.01 S1F, site 1 binding factor of spinach 0.79 997 1013 (−) 1.000 0.821 gctgATGGgaagaagaa rps1 promoter SEQ_22 P$GTBX/SBF1.01 SBF-1 0.87 1016 1032 (+) 1.000 0.875 tttctttTTAAaaaatt SEQ_22 P$STKM/STK.01 Storekeeper (STK), plant specific 0.85 1021 1035 (+) 1.000 0.881 tttTAAAaaattgaa DNA binding protein important for tuber-specific and sucrose-inducible gene expression SEQ_22 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 1039 1049 (−) 1.000 0.852 aAAAAgttaaa SEQ_22 P$GTBX/SBF1.01 SBF-1 0.87 1042 1058 (−) 1.000 0.874 gaaatttTTAAaaagtt SEQ_22 P$OPAQ/O2.01 Opaque-2 regulatory protein 0.87 1066 1082 (−) 1.000 0.898 tccataataTCATctga SEQ_22 P$AGP1/AGP1.01 AG-motif binding protein 1 0.91 1080 1090 (−) 1.000 0.915 tgaGATCttcc SEQ_22 P$AGP1/AGP1.01 AG-motif binding protein 1 0.91 1081 1091 (+) 1.000 0.911 gaaGATCtcaa SEQ_22 P$L1BX/PDF2.01 Protodermal factor 2 0.85 1090 1106 (+) 1.000 0.897 aagagtTAAAtgtatcc SEQ_22 P$MYBS/OSMYBS.01 Rice MYB proteins with single DNA 0.82 1097 1113 (+) 1.000 0.897 aaatgTATCcatcttgg binding domains, binding to the amylase element (TATCCA) SEQ_22 P$L1BX/PDF2.01 Protodermal factor 2 0.85 1111 1127 (−) 0.848 0.899 ccggttTTAAtgcccca SEQ_22 P$L1BX/ATML1.02 Arabidopsis thaliana meristem layer 1 0.76 1113 1129 (+) 1.000 0.854 gggCATTaaaaccggtg SEQ_22 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 1115 1125 (−) 1.000 1.000 ggtttTAATgc SEQ_22 P$IBOX/GATA.01 Class I GATA factors 0.93 1133 1149 (+) 1.000 0.949 gggatGATAaatacaga SEQ_22 P$L1BX/PDF2.01 Protodermal factor 2 0.85 1134 1150 (+) 1.000 0.855 ggatgaTAAAtacagac SEQ_22 P$ROOT/RHE.02 Root hair-specific element with a 3- 0.77 1172 1196 (+) 1.000 0.804 gtaattcatatttatCACGttgcta nucleotid spacer between left part (LP) and right part (RP) SEQ_22 P$NACF/TANAC69.01 Wheat NACdomain DNA binding 0.68 1176 1198 (+) 0.895 0.684 ttcatatttatCACGttgctaaa factor SEQ_22 P$GBOX/HBP1B.01 Wheat bZIP transcription factor 0.83 1179 1199 (−) 1.000 0.835 ttttagcaACGTgataaatat HBP1B (histone gene binding protein 1b) SEQ_22 P$MYCL/MYCRS.01 Myc recognition sequences 0.93 1180 1198 (−) 1.000 0.967 tttagcaACGTgataaata SEQ_22 P$ABRE/ABRE.01 ABA response elements 0.82 1181 1197 (+) 1.000 0.826 atttatcACGTtgctaa SEQ_22 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription 0.81 1182 1198 (+) 0.951 0.830 tttatcACGTtgctaaa factors that belong to the group of Opaque-2 like proteins Storekeeper (STK), plant specific SEQ_22 P$STKM/STK.01 DNA binding protein important for 0.85 1192 1206 (+) 1.000 0.888 tgcTAAAaaaattat tuber-specific and sucrose-inducible gene expression SEQ_22 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 1223 1249 (+) 0.750 0.609 acaaaatCAATtaaagagaaagaaaga element about 100 bp up- stream of the TSS in legumin genes SEQ_22 P$DOFF/DOF1.01 Dof1/MNB1a - single zinc finger 0.98 1227 1243 (+) 1.000 0.991 aatcaattAAAGagaaa transcription factor SEQ_22 P$GAGA/GAGABP.01 (GA)n/(CT)n binding proteins (GBP, 0.75 1231 1255 (+) 1.000 0.790 aattaaAGAGaaagaaagaaacgca soybean; BBR, barley) SEQ_22 P$NCS2/NCS2.01 Nodulin consensus sequence 2 0.79 1232 1246 (−) 1.000 0.808 ttctttCTCTttaat SEQ_22 P$OCSE/OCSL.01 OCS-like elements 0.69 1283 1303 (+) 0.769 0.735 taaaataaaaattaACGCatg SEQ_22 P$SEF4/SEF4.01 Soybean embryo factor 4 0.98 1285 1295 (−) 1.000 0.983 aaTTTTtattt SEQ_22 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription 0.81 1294 1310 (−) 1.000 0.825 cattcaACATgcgttaa factors that belong to the group of Opaque-2 like proteins SEQ_22 P$IDRE/IDE1.01 Iron-deficiency-responsive element 1 0.77 1297 1311 (+) 1.000 0.773 acGCATgttgaatgc SEQ_22 P$GBOX/BZIP910.02 bZIP transcription factor from Antirrhinum 0.84 1300 1320 (+) 1.000 0.864 catgttgaatgcTGACatgtc majus SEQ_22 P$GBOX/BZIP910.01 bZIP transcription factor from Antirrhinum 0.77 1306 1326 (+) 1.000 0.784 gaatgcTGACatgtcagtatg majus SEQ_22 P$ABRE/ABF1.03 ABA (abscisic acid) inducible transcriptional 0.82 1307 1323 (+) 0.750 0.829 aatgctgaCATGtcagt activator SEQ_22 P$OPAQ/O2.01 Opaque-2 regulatory protein 0.87 1307 1323 (−) 0.794 0.901 actgacatgTCAGcatt SEQ_22 P$GTBX/S1F.01 S1F, site 1 binding factor of spinach 0.79 1319 1335 (−) 1.000 0.818 attcATGGacatactga rps1 promoter SEQ_22 P$ROOT/RHE.01 Root hair-specific element with a 2- 0.77 1322 1346 (+) 1.000 0.844 gtatgtccatgaatcCACGtatcaa nucleotid spacer between left part (LP) and right part (RP) SEQ_22 P$GBOX/GBF1.01 bZIP protein G-Box binding factor 1 0.94 1329 1349 (−) 1.000 0.956 cgcttgatACGTggattcatg SEQ_22 P$GBOX/HBP1A.01 HBP-1a, suggested to be involved in 0.88 1330 1350 (+) 1.000 0.899 atgaatcCACGtatcaagcgc the cell cycle-dependent expression SEQ_22 P$ABRE/ABRE.01 ABA response elements 0.82 1332 1348 (−) 1.000 0.866 gcttgatACGTggattc SEQ_22 P$L1BX/ATML1.01 L1-specific homeodomain protein 0.82 1370 1386 (+) 1.000 0.889 tctttcTAAAtgaaaac ATML1 (A. thaliana meristem layer 1) SEQ_22 P$GAPB/GAP.01 Cis-element in the GAPDH promoters 0.88 1375 1389 (+) 1.000 0.958 ctaaATGAaaacaac conferring light inducibility SEQ_22 P$URNA/USE.01 Upstream sequence elements in the 0.75 1387 1403 (+) 0.750 0.781 aacttcACACatcacaa promoters of U-snRNA genes of higher plants SEQ_22 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 1396 1412 (−) 0.817 0.771 tattgtTTGTtgtgatg SEQ_22 P$DPBF/DPBF.01 bZIP factors DPBF-1 and 2 (Dc3 0.89 1413 1423 (+) 1.000 0.898 cACACaagacc promoter binding factor-1 and 2) SEQ_22 P$GAGA/BPC.01 Basic pentacysteine proteins 1.00 1413 1437 (−) 1.000 1.000 aacgagAGAGagggggtcttgtgtg SEQ_22 P$GAGA/GAGABP.01 (GA)n/(CT)n binding proteins (GBP, 0.75 1423 1447 (−) 0.750 0.805 gcagagAGACaacgagagagagggg soybean; BBR, barley) SEQ_22 P$GAGA/BPC.01 Basic pentacysteine proteins 1.00 1425 1449 (−) 1.000 1.000 tggcagAGAGacaacgagagagagg SEQ_22 P$PREM/MGPROTORE.01 Promoter elements involved in 0.77 1447 1477 (+) 1.000 0.794 ccagCGACcaaatcgaagcttgagaagaaca MgProto (Mg-protoporphyrin IX) and light-mediated induction

TABLE 7 cis-regulatory elements of SEQ ID NO: 25 Seq. Opt. Start End Core Matrix name Family/matrix Further Information thresh. pos. pos. Strand sim. sim. Sequence SEQ_25 P$IBOX/GATA.01 Class I GATA factors 0.93 27 43 (+) 1.000 0.946 aaaaaGATAaccacccc SEQ_25 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 27 37 (+) 1.000 0.948 aAAAAgataac SEQ_25 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 29 45 (−) 1.000 0.980 agggggtgGTTAtcttt SEQ_25 P$SALT/ALFIN1.02 Zinc-finger protein in alfalfa roots, 0.95 34 48 (−) 1.000 0.977 gctagggGGTGgtta regulates salt tolerance SEQ_25 P$SUCB/SUCROSE.01 Sequence motif from the promoters 0.81 57 75 (+) 1.000 0.852 ccAAATcataactatcaga of different sugar-responsive genes SEQ_25 P$AHBP/ATHB9.01 HD-ZIP class III protein ATHB9 0.77 58 68 (−) 1.000 0.772 gttATGAtttg SEQ_25 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 60 76 (−) 1.000 0.808 ttctgaTAGTtatgatt SEQ_25 P$IBOX/GATA.01 Class I GATA factors 0.93 76 92 (+) 1.000 0.975 acaaaGATAaaaagccc SEQ_25 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription 0.99 78 94 (+) 1.000 0.996 aaagataaAAAGcccga factor SEQ_25 P$SBPD/SBP.01 SQUA promoter binding proteins 0.88 99 115 (−) 1.000 0.882 ctatgGTACaacatggt SEQ_25 P$NCS3/NCS3.01 Nodulin consensus sequence 3 0.89 119 129 (+) 1.000 0.893 caCACCctcta SEQ_25 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 122 148 (−) 0.750 0.626 ttcataaCTATgtatgagatagagggt element about 100 by up- stream of the TSS in legumin genes SEQ_25 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 133 149 (+) 1.000 0.778 catacaTAGTtatgaat SEQ_25 P$MIIG/P_ACT.01 Maize activator P of flavonoid bio- 0.93 150 164 (−) 1.000 0.977 ttacGGTAggtttca synthetic genes SEQ_25 P$MYBL/ATMYB77.01 R2R3-type myb-like transcription 0.87 172 188 (−) 1.000 0.909 ttgtggCGGTtcctgct factor (I-type binding site) SEQ_25 P$DREB/HVDRF1.01 H. vulgare dehydration-response 0.89 175 189 (+) 1.000 0.953 aggaACCGccacaat factor 1 SEQ_25 P$GCCF/ERE_JERE.01 Ethylene-responsive elements (ERE) 0.85 175 187 (+) 1.000 0.865 aggaacCGCCaca and jasmonate- and elicitor- responsive elements (JERE) SEQ_25 P$MYBS/TAMYB80.01 MYB protein from wheat 0.83 191 207 (−) 1.000 0.874 gcttATATtcccctcac SEQ_25 P$TEFB/TEF1.01 TEF cis acting elements in both RNA 0.76 192 212 (+) 1.000 0.796 tgAGGGgaatataagccaaag polymerase II-dependent promoters and rDNA spacer sequences SEQ_25 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 206 232 (+) 0.750 0.649 gccaaagCCCTgcaattttcagtgaga element about 100 by up- stream of the TSS in legumin genes SEQ_25 P$TEFB/TEF1.01 TEF cis acting elements in both RNA 0.76 230 250 (+) 0.956 0.795 agAAGGgtaagattattaaag polymerase II-dependent promoters and rDNA spacer sequences SEQ_25 P$AHBP/HAHB4.01 Sunflower homeodomain leucine- 0.87 238 248 (+) 1.000 0.903 aagattATTAa zipper protein Hahb-4 SEQ_25 P$DOFF/DOF1.01 Dof1/MNB1a - single zinc finger 0.98 239 255 (+) 1.000 0.980 agattattAAAGgcagc transcription factor SEQ_25 P$L1BX/ATML1.01 L1-specific homeodomain protein 0.82 240 256 (+) 1.000 0.835 gattatTAAAggcagcc ATML1 (A. thaliana meristem layer 1) SEQ_25 P$DREB/CRT_DRE.01 C-repeat/dehydration response element 0.89 262 276 (−) 1.000 0.968 ctttgCCGAcattgt SEQ_25 P$URNA/USE.01 Upstream sequence elements in the 0.75 284 300 (−) 1.000 0.858 aatgtcCCACctcgaat promoters of U-snRNA genes of higher plants SEQ_25 P$GTBX/SBF1.01 SBF-1 0.87 292 308 (+) 1.000 0.908 tgggacaTTAAatttaa SEQ_25 P$L1BX/HDG9.01 Homeodomain glabrous 9 0.77 292 308 (−) 0.796 0.792 ttaaatTTAAtgtccca SEQ_25 P$L1BX/PDF2.01 Protodermal factor 2 0.85 294 310 (+) 1.000 0.851 ggacatTAAAtttaaaa SEQ_25 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 296 306 (−) 1.000 1.000 aaattTAATgt SEQ_25 P$GTBX/SBF1.01 SBF-1 0.87 298 314 (+) 1.000 0.875 attaaatTTAAaaagaa SEQ_25 P$GTBX/SBF1.01 SBF-1 0.87 299 315 (−) 1.000 0.885 cttctttTTAAatttaa SEQ_25 P$IDRE/IDE1.01 Iron-deficiency-responsive element 1 0.77 325 339 (−) 0.809 0.806 aaGCTTgctactttc SEQ_25 P$AGP1/AGP1.01 AG-motif binding protein 1 0.91 376 386 (+) 1.000 0.912 caaGATCttcc SEQ_25 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 383 393 (+) 1.000 0.963 ttcctTAATcc SEQ_25 P$GTBX/S1F.01 S1F, site 1 binding factor of spinach 0.79 383 399 (−) 1.000 0.810 tgttATGGattaaggaa rps1 promoter SEQ_25 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain 0.82 387 407 (+) 1.000 0.865 ttaatCCATaacaagaagtcc protein AGAMOUS-like 2 SEQ_25 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 435 445 (+) 0.829 0.940 acaATGAttct SEQ_25 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 435 445 (−) 0.936 0.941 agaATCAttgt SEQ_25 P$SUCB/SUCROSE.01 Sequence motif from the promoters 0.81 444 462 (+) 1.000 0.875 ctAAATcatacatattacc of different sugar-responsive genes SEQ_25 P$MADS/AGL15.01 AGL15, Arabidopsis MADS-domain 0.79 487 507 (−) 0.850 0.804 tttTGCTacacctggtagtag protein AGAMOUS-like 15 SEQ_25 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), 0.95 487 505 (−) 0.954 0.966 ttgctACACctggtagtag AtMYC2 (rd22BP1) SEQ_25 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain 0.82 488 508 (+) 0.869 0.855 tactaCCAGgtgtagcaaaat protein AGAMOUS-like 2 SEQ_25 P$L1BX/PDF2.01 Protodermal factor 2 0.85 510 526 (+) 1.000 0.880 cccgatTAAAttcataa SEQ_25 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 512 522 (−) 1.000 0.963 gaattTAATcg SEQ_25 P$RAV5/RAV1-5.01 5′-part of bipartite RAV1 binding site, 0.96 537 547 (−) 1.000 0.974 agcAACAaaat interacting with AP2 domain SEQ_25 P$CARM/CARICH.01 CA-rich element 0.78 548 566 (+) 1.000 0.837 accttcaAACAacagatgc SEQ_25 P$MYCL/ICE.01 ICE (inducer of CBF expression 1), 0.95 554 572 (+) 0.863 0.954 aaacaACAGatgctcgcaa AtMYC2 (rd22BP1) SEQ_25 P$RAV5/RAV1-5.01 5′-part of bipartite RAV1 binding site, 0.96 555 565 (+) 1.000 0.961 aacAACAgatg interacting with AP2 domain SEQ_25 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 596 612 (+) 1.000 0.771 tctagcTAGTaacgacc SEQ_25 P$MYBL/NTMYBAS1.01 Anther-specific myb gene from tobacco 0.96 600 616 (−) 1.000 0.975 gttaggtcGTTActagc SEQ_25 P$MYBL/GAMYB.01 GA-regulated myb gene from barley 0.91 608 624 (−) 1.000 0.927 atagtgttGTTAggtcg SEQ_25 P$MIIG/PALBOXL.01 Cis-acting element conserved in 0.80 622 636 (−) 1.000 0.812 atctgtttGGTGata various PAL and 4CL promoters SEQ_25 P$CARM/CARICH.01 CA-rich element 0.78 623 641 (+) 1.000 0.788 atcaccaAACAgataaaca SEQ_25 P$MYBL/CARE.01 CAACTC regulatory elements, GA- 0.83 646 662 (+) 1.000 0.834 tctagcgAGTTccagca inducible SEQ_25 P$CE1F/ABI4.01 ABA insensitive protein 4 (ABI4) 0.87 680 692 (+) 1.000 0.900 tcgcCACCgacga SEQ_25 P$DREB/CRT_DRE.01 C-repeat/dehydration response element 0.89 681 695 (+) 1.000 0.914 cgccaCCGAcgatta SEQ_25 P$PREM/MGPROTORE.01 Promoter elements involved in 0.77 683 713 (+) 1.000 0.774 ccacCGACgattatcgattcactaaagctac MgProto (Mg-protoporphyrin IX) and light-mediated induction SEQ_25 P$DOFF/DOF1.01 Dof1/MNB1a - single zinc finger 0.98 698 714 (+) 1.000 1.000 gattcactAAAGctaca transcription factor SEQ_25 P$CGCG/ATSR1.01 Arabidopsis thaliana signal- 0.84 714 730 (−) 1.000 0.865 ccaCGCGtgtacttgtt responsive gene1, Ca2+/calmodulin binding protein homolog to NtER1 (tobacco early ethylene-responsive gene) SEQ_25 P$CE3S/CE3.01 Coupling element 3 (CE3), non- 0.77 715 733 (−) 1.000 0.787 tatccaCGCGtgtacttgt ACGT ABRE SEQ_25 P$CGCG/ATSR1.01 Arabidopsis thaliana signal- 0.84 721 737 (+) 1.000 0.870 acaCGCGtggatagtgg responsive gene1, Ca2+/calmodulin binding protein homolog to NtER1 (tobacco early ethylene-responsive gene) SEQ_25 P$MYBS/MYBST1.01 MybSt1 (Myb Solanum tuberosum 1) 0.90 722 738 (−) 1.000 0.936 tccactATCCacgcgtg with a single myb repeat SEQ_25 P$HOCT/HOCT.01 Octamer motif found in plant histone 0.76 723 739 (−) 1.000 0.768 ttccactATCCacgcgt H3 and H4 genes SEQ_25 P$MYBS/ZMMRP1.01 Zea mays MYB-related protein 1 0.79 732 748 (−) 0.777 0.852 atttctcTATTccacta (transfer cell specific) SEQ_25 P$NCS2/NCS2.01 Nodulin consensus sequence 2 0.79 746 760 (+) 0.750 0.803 aattgcCTGTtcaac SEQ_25 P$MSAE/MSA.01 M-phase-specific activators 0.80 753 767 (+) 1.000 0.875 tgttcAACGgggaga (NtmybA1, NtmybA2, NtmybB) SEQ_25 P$PREM/MGPROTORE.01 Promoter elements involved in 0.77 786 816 (+) 1.000 0.801 atagCGACaaggaggaggagcgatattgcta MgProto (Mg-protoporphyrin IX) and light-mediated induction SEQ_25 P$IDDF/ID1.01 Maize INDETERMINATE1 zinc finger 0.92 787 799 (−) 1.000 0.921 ctccTTGTcgcta protein SEQ_25 P$MYBS/MYBST1.01 MybSt1 (Myb Solanum tuberosum 1) 0.90 810 826 (+) 1.000 0.928 attgctATCCggaaagt with a single myb repeat SEQ_25 P$DREB/HVDRF1.01 H. vulgare dehydration-response 0.89 842 856 (+) 0.826 0.914 actcGCCGccatata factor 1 SEQ_25 P$GARP/ARR10.01 Type-B response regulator (ARR10), 0.97 857 865 (−) 1.000 0.970 AGATccttg member of the GARP-family of plant myb-related DNA binding motifs SEQ_25 P$SALT/ALFIN1.01 Zinc-finger protein in alfalfa roots, 0.93 872 886 (−) 1.000 0.930 ccttgGTGGcgccgt regulates salt tolerance SEQ_25 P$PREM/MGPROTORE.01 Promoter elements involved in 0.77 888 918 (−) 1.000 0.774 actaCGACggcgatgagggtgaccattcgag MgProto (Mg-protoporphyrin IX) and light-mediated induction SEQ_25 P$NCS3/NCS3.01 Nodulin consensus sequence 3 0.89 896 906 (+) 1.000 0.947 gtCACCctcat SEQ_25 P$AHBP/ATHB9.01 HD-ZIP class III protein ATHB9 0.77 920 930 (+) 0.750 0.773 gtaTTGAtctc SEQ_25 P$GARP/ARR10.01 Type-B response regulator (ARR10), 0.97 941 949 (+) 1.000 0.973 AGATcctgg member of the GARP-family of plant myb-related DNA binding motifs SEQ_25 P$ABRE/ABF1.03 ABA (abscisic acid) inducible transcriptional 0.82 952 968 (−) 1.000 0.833 ggcgaggcCGTGgctca activator SEQ_25 P$L1BX/ATML1.02 Arabidopsis thaliana meristem layer 1 0.76 987 1003 (−) 1.000 0.791 aggCATTcaaatctggc SEQ_25 P$L1BX/PDF2.01 Protodermal factor 2 0.85 989 1005 (+) 0.787 0.854 cagattTGAAtgcctcc SEQ_25 P$HEAT/HSE.01 Heat shock element 0.81 1015 1029 (−) 1.000 0.822 gcatatctccAGAAt SEQ_25 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 1076 1092 (+) 0.817 0.775 gagagaTTGTtgctttc SEQ_25 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 1077 1091 (−) 1.000 0.893 aaagcaacAATCtct SEQ_25 P$GTBX/GT3A.01 Trihelix DNA-binding factor GT-3a 0.83 1107 1123 (+) 1.000 0.847 atcagtGTTActtcgat SEQ_25 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 1126 1140 (−) 1.000 0.898 gcaaacagAATCtca SEQ_25 P$GAGA/BPC.01 Basic pentacysteine proteins 1.00 1128 1152 (−) 1.000 1.000 cgagagAGAGaggcaaacagaatct SEQ_25 P$GAGA/BPC.01 Basic pentacysteine proteins 1.00 1130 1154 (−) 1.000 1.000 aacgagAGAGagaggcaaacagaat SEQ_25 P$HEAT/HSE.01 Heat shock element 0.81 1130 1144 (−) 1.000 0.855 agaggcaaacAGAAt SEQ_25 P$GAGA/GAGABP.01 (GA)n/(CT)n binding proteins (GBP, 0.75 1134 1158 (−) 0.750 0.760 ccagaaCGAGagagagaggcaaaca soybean; BBR, barley) SEQ_25 P$GAGA/GAGABP.01 (GA)n/(CT)n binding proteins (GBP, 0.75 1138 1162 (−) 0.750 0.757 gaaaccAGAAcgagagagagaggca soybean; BBR, barley) SEQ_25 P$PSRE/GAAA.01 GAAA motif involved in pollen specific 0.83 1164 1180 (+) 1.000 0.843 ctgtaGAAAaacttttt transcriptional activation SEQ_25 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 1170 1180 (−) 1.000 0.853 aAAAAgttttt SEQ_25 P$PSRE/GAAA.01 GAAA motif involved in pollen specific 0.83 1185 1201 (−) 1.000 0.838 ctaaaGAAAatgttcgc transcriptional activation SEQ_25 P$DOFF/DOF1.01 Dof1/MNB1a - single zinc finger 0.98 1191 1207 (−) 1.000 0.984 ttgggcctAAAGaaaat transcription factor SEQ_25 P$TCPF/ATTCP20.01 TCP class I transcription factor 0.94 1198 1210 (+) 1.000 0.968 ttagGCCCaaagt (Arabidopsis) SEQ_25 P$TCPF/ATTCP20.01 TCP class I transcription factor 0.94 1208 1220 (−) 1.000 0.943 aaaaGCCCaaact (Arabidopsis) SEQ_25 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription 0.99 1211 1227 (−) 1.000 0.994 cgttattaAAAGcccaa factor SEQ_25 P$L1BX/HDG9.01 Homeodomain glabrous 9 0.77 1211 1227 (−) 1.000 0.818 cgttatTAAAagcccaa SEQ_25 P$GTBX/SBF1.01 SBF-1 0.87 1213 1229 (−) 1.000 0.927 ggcgttaTTAAaagccc SEQ_25 P$DOFF/DOF3.01 Dof3 - single zinc finger transcription 0.99 1223 1239 (+) 1.000 0.995 taacgcctAAAGcccaa factor SEQ_25 P$CAAT/CAAT.01 CCAAT-box in plant promoters 0.97 1250 1258 (−) 1.000 0.979 acCCAAtaa SEQ_25 P$TEFB/TEF1.01 TEF cis acting elements in both RNA 0.76 1288 1308 (+) 0.956 0.767 atAAGGggaatctatttattt polymerase II-dependent promoters and rDNA spacer sequences SEQ_25 P$MADS/SQUA.01 MADS-box protein SQUAMOSA 0.90 1294 1314 (+) 1.000 0.917 ggaatctATTTatttaattgt SEQ_25 P$STKM/STK.01 Storekeeper (STK), plant specific 0.85 1298 1312 (−) 1.000 0.859 aatTAAAtaaataga DNA binding protein important for tuber-specific and sucrose-inducible gene expression SEQ_25 P$L1BX/HDG9.01 Homeodomain glabrous 9 0.77 1299 1315 (−) 1.000 0.791 aacaatTAAAtaaatag SEQ_25 P$HMGF/HMG_IY.01 High mobility group I/Y-like proteins 0.89 1300 1314 (+) 1.000 0.907 tattTATTtaattgt SEQ_25 P$GTBX/SBF1.01 SBF-1 0.87 1307 1323 (+) 1.000 0.872 ttaattgTTAAtcattc SEQ_25 P$AHBP/WUS.01 Homeodomain protein WUSCHEL 0.94 1310 1320 (+) 1.000 0.963 attgtTAATca SEQ_25 P$OCSE/OCSL.01 OCS-like elements 0.69 1310 1330 (+) 1.000 0.712 attgttaatcattcACGTtga SEQ_25 P$LEGB/LEGB.01 Legumin box, highly conserved sequence 0.59 1311 1337 (+) 0.750 0.625 ttgttaaTCATtcacgttgaccattga element about 100 bp up- stream of the TSS in legumin genes SEQ_25 P$AHBP/ATHB5.01 HDZip class I protein ATHB5 0.89 1314 1324 (+) 0.936 0.939 ttaATCAttca SEQ_25 P$AHBP/HAHB4.01 Sunflower homeodomain leucine- 0.87 1314 1324 (−) 1.000 0.945 tgaatgATTAa zipper protein Hahb-4 SEQ_25 P$GBOX/HBP1B.01 Wheat bZIP transcription factor 0.83 1315 1335 (−) 1.000 0.834 aatggtcaACGTgaatgatta HBP1B (histone gene binding protein 1b) SEQ_25 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription 0.81 1317 1333 (−) 0.951 0.842 tggtcaACGTgaatgat factors that belong to the group of Opaque-2 like proteins SEQ_25 P$WBXF/ERE.01 Elicitor response element 0.89 1322 1338 (+) 1.000 0.917 tcacgtTGACcattgaa SEQ_25 P$HEAT/HSE.01 Heat shock element 0.81 1328 1342 (+) 1.000 0.826 tgaccattgaAGAAc SEQ_25 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 1373 1383 (−) 0.804 0.896 cAAAGgatcaa SEQ_25 P$MADS/AGL2.01 AGL2, Arabidopsis MADS-domain 0.82 1408 1428 (+) 0.869 0.838 tgtctCCAGctctagtaatga protein AGAMOUS-like 2 SEQ_25 P$MYBL/MYBPH3.02 Myb-like protein of Petunia hybrida 0.76 1427 1443 (+) 1.000 0.785 gacaatTAGTttagttt SEQ_25 P$STKM/STK.01 Storekeeper (STK), plant specific 0.85 1428 1442 (−) 1.000 0.864 aacTAAActaattgt DNA binding protein important for tuber-specific and sucrose-inducible gene expression SEQ_25 P$SPF1/SP8BF.01 DNA-binding protein of sweet potato 0.87 1459 1471 (−) 1.000 0.989 aaTACTattacac that binds to the SP8a (ACTGTGTA) and SP8b (TACTATT) sequences of sporamin and beta-amylase genes SEQ_25 P$GTBX/SBF1.01 SBF-1 0.87 1495 1511 (−) 1.000 0.886 aaaattgTTAAataata SEQ_25 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 1497 1513 (−) 1.000 0.809 agaaaattGTTAaataa SEQ_25 P$E2FF/E2F.01 E2F class I sites 0.82 1527 1541 (+) 1.000 0.832 ttttTTCCagcaacg SEQ_25 P$GTBX/SBF1.01 SBF-1 0.87 1548 1564 (+) 1.000 0.874 tgaatttTTAAaacttg SEQ_25 P$SUCB/SUCROSE.01 Sequence motif from the promoters 0.81 1552 1570 (−) 0.750 0.832 aaATATcaagttttaaaaa of different sugar-responsive genes SEQ_25 P$CCAF/CCA1.01 Circadian clock associated 1 0.85 1568 1582 (−) 1.000 0.854 ttaaaaaaAATCaaa SEQ_25 P$AHBP/BLR.01 Transcriptional repressor BELL- 0.90 1581 1591 (−) 1.000 0.976 taaATTActtt RINGER SEQ_25 P$GTBX/SBF1.01 SBF-1 0.87 1583 1599 (−) 1.000 0.900 tttgcatTTAAattact SEQ_25 P$L1BX/ATML1.01 L1-specific homeodomain protein 0.82 1584 1600 (+) 1.000 0.918 gtaattTAAAtgcaaaa ATML1 (A. thaliana meristem layer 1) SEQ_25 P$NCS1/NCS1.01 Nodulin consensus sequence 1 0.85 1606 1616 (+) 0.878 0.862 cAAATgatatt SEQ_25 P$MYBL/CARE.01 CAACTC regulatory elements, GA- 0.83 1625 1641 (+) 1.000 0.834 actcaagAGTTgtgtga inducible SEQ_25 P$NCS2/NCS2.01 Nodulin consensus sequence 2 0.79 1653 1667 (−) 1.000 0.809 acttgcCTCTtgccc SEQ_25 P$TEFB/TEF1.01 TEF cis acting elements in both RNA 0.76 1665 1685 (−) 0.956 0.816 agAAGGatacaccagtgcact polymerase II-dependent promoters and rDNA spacer sequences SEQ_25 P$MYBS/HVMCB1.01 Hordeum vulgare Myb-related CAB- 0.93 1672 1688 (+) 1.000 0.933 tggtgtATCCttctcgg promoter-binding protein 1 SEQ_25 P$GTBX/SBF1.01 SBF-1 0.87 1686 1702 (+) 1.000 0.878 cggggcgTTAAaaccgt SEQ_25 P$NCS3/NCS3.01 Nodulin consensus sequence 3 0.89 1702 1712 (−) 1.000 0.965 gtCACCttcca SEQ_25 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 1704 1720 (−) 0.750 0.832 attaaacgGTCAccttc SEQ_25 P$MSAE/MSA.01 M-phase-specific activators 0.80 1707 1721 (−) 1.000 0.897 tattaAACGgtcacc (NtmybA1, NtmybA2, NtmybB) SEQ_25 P$L1BX/HDG9.01 Homeodomain glabrous 9 0.77 1708 1724 (−) 1.000 0.791 atgtatTAAAcggtcac SEQ_25 P$OPAQ/O2_GCN4.01 Recognition site for BZIP transcription 0.81 1716 1732 (−) 1.000 0.834 agatagACATgtattaa factors that belong to the group of Opaque-2 like proteins SEQ_25 P$LREM/ATCTA.01 Motif involved in carotenoid and 0.85 1727 1737 (+) 1.000 0.853 ctATCTattat tocopherol biosynthesis and in the expression of photosynthesis-related genes SEQ_25 P$ROOT/RHE.02 Root hair-specific element with a 3- 0.77 1747 1771 (+) 1.000 0.774 gtacttcatagctatCACGttgctc nucleotid spacer between left part (LP) and right part (RP) SEQ_25 P$GBOX/HBP1B.01 Wheat bZIP transcription factor 0.83 1754 1774 (−) 1.000 0.843 gaggagcaACGTgatagctat HBP1B (histone gene binding protein 1b) SEQ_25 P$OCSE/OCSL.01 OCS-like elements 0.69 1760 1780 (−) 1.000 0.700 atttgtgaggagcaACGTgat SEQ_25 P$SUCB/SUCROSE.01 Sequence motif from the promoters 0.81 1799 1817 (+) 1.000 0.911 gcAAATcaattttataaag of different sugar-responsive genes SEQ_25 P$TBPF/TATA.01 Plant TATA box 0.88 1807 1821 (+) 1.000 0.910 atttTATAaagacgc SEQ_25 P$GTBX/GT3A.01 Trihelix DNA-binding factor GT-3a 0.83 1820 1836 (−) 1.000 0.865 tcagctGTTActcatgc SEQ_25 P$MYBL/MYBPH3.01 Myb-like protein of Petunia hybrida 0.80 1822 1838 (−) 1.000 0.823 attcagctGTTActcat SEQ_25 P$DOFF/PBF.01 PBF (MPBF) 0.97 1834 1850 (+) 1.000 0.988 tgaataaaAAAGagagg SEQ_25 P$IDRE/IDE1.01 Iron-deficiency-responsive element 1 0.77 1856 1870 (+) 1.000 0.777 acGCATgttgattgc SEQ_25 P$GBOX/BZIP910.02 bZIP transcription factor from Antirrhinum 0.84 1859 1879 (+) 1.000 0.962 catgttgattgcTGACgtgtc majus SEQ_25 P$OCSE/OCSL.01 OCS-like elements 0.69 1859 1879 (+) 1.000 0.751 catgttgattgctgACGTgtc SEQ_25 P$GBOX/HBP1B.01 Wheat bZIP transcription factor 0.83 1864 1884 (−) 1.000 0.973 ctacggacACGTcagcaatca HBP1B (histone gene binding protein 1b) SEQ_25 P$GBOX/GBF1.01 bZIP protein G-Box binding factor 1 0.94 1865 1885 (+) 1.000 0.999 gattgctgACGTgtccgtagg SEQ_25 P$ABRE/ABF1.03 ABA (abscisic acid) inducible transcriptional 0.82 1866 1882 (+) 1.000 0.960 attgctgaCGTGtccgt activator SEQ_25 P$OPAQ/RITA1.01 Rice transcription activator-1 (RITA), 0.95 1866 1882 (−) 1.000 0.956 acggacACGTcagcaat basic leucin zipper protein, highly expressed during seed development SEQ_25 P$OPAQ/RITA1.01 Rice transcription activator-1 (RITA), 0.95 1867 1883 (+) 1.000 0.958 ttgctgACGTgtccgta basic leucin zipper protein, highly expressed during seed development SEQ_25 P$EINL/TEIL.01 TEIL (tobacco EIN3-like) 0.92 1882 1890 (−) 0.863 0.966 aTGGAccta SEQ_25 P$GAGA/BPC.01 Basic pentacysteine proteins 1.00 1937 1961 (−) 1.000 1.000 ctcgagAGAGaggggacttatgatg

In the following table 9, cis-regulatory element families commonly found in at least 13 of the listed expression control sequences of the present invention are shown. Table 8 describes the cis-regulatory element families.

TABLE 8 Description of cis-regulatory elements families commonly found in expression control sequences of the present invention. TF-Family Description O$VTBP Vertebrate TATA binding protein vector P$AHBP Arabidopsis homeobox protein P$DOFF DNA binding with one finger (DOF) P$GTBX GT-box elements P$IBOX Plant I-Box sites P$L1BX L1 box, motif for L1 layer-specific expression P$LREM Light responsive element motif, not modulated by different light qualities P$MYBL MYB-like proteins P$MYBS MYB proteins with single DNA binding repeat P$NCS1 Nodulin consensus sequence 1 P$CCAF Circadian control factors P$GBOX Plant G-box/C-box bZIP proteins P$IDDF D domain factors, the ID domain includes a cluster of three different types of zinc fingers seperated from a fourth C2H2 finger by a long spacer P$MADS floral determination P$MYCL Myc-like basic helix-loop-helix binding factors P$OPAQ Opaque-2 like transcriptional activators P$SPF1 Sweet potato DNA-binding factor with two WRKY-domains P$SUCB Sucrose box P$WBXF W Box family O$INRE Core promoter initiator elements O$PTBP Plant TATA binding protein vector P$ABRE ABA response elements P$LEGB Legumin Box family P$NACF Plant specific NAC [NAM (no apical meristem), ATAF172, CUC2 (cup-shaped cotyledons 2)] transcription factors P$OCSE Enhancer element first identified in the promoter of the octopine synthase gene (OCS) of the Agrobacterium tumefaciens T-DNA

TABLE 9 Cis-regulatory elements families commonly found in expression control sequences in at least 13 of the listed expression control sequences of the present invention. No of occurences p- p- p- p- p- BnSCP p- p- p- BnSETL- BnSETL- BnSCT2- BnSCT2- p- (SEQ- BnGRPL BnCRU4 BnMYR var1 var2 var1 var2 BnMDP TF- ID (SEQ-ID (SEQ-ID (SEQ- (SEQ-ID (SEQ-ID (SEQ-ID (SEQ-ID (SEQ- Family 70) 1) 77) ID 85) 22) 25) 14) 16) ID 95) O$VTBP 7 18 17 39 6 10 5 8 19 P$AHBP 16 8 22 25 6 12 7 9 15 P$DOFF 14 8 11 9 2 7 5 5 4 P$GTBX 12 13 10 16 10 12 5 1 7 P$IBOX 4 4 4 2 4 2 2 2 7 P$L1BX 9 7 11 18 10 10 2 3 10 P$LREM 6 4 4 6 3 1 3 3 5 P$MYBL 9 11 12 7 16 14 9 7 9 P$MYBS 9 3 8 2 4 5 2 2 10 P$NCS1 2 2 4 3 3 4 2 2 2 P$CCAF 4 4 4 2 3 3 1 1 3 P$GBOX 9 3 12 2 5 5 7 5 3 P$IDDF 3 2 1 1 1 1 4 5 3 P$MADS 6 9 9 2 0 5 2 2 7 P$MYCL 1 0 6 3 5 2 3 3 3 P$OPAQ 9 5 7 3 6 4 1 2 4 P$SPF1 2 6 1 5 1 1 2 2 2 P$SUCB 6 4 3 7 1 4 2 3 0 P$WBXF 2 3 1 0 1 1 3 2 1 O$INRE 5 1 1 2 1 4 0 0 1 O$PTBP 2 7 8 24 1 1 0 1 5 P$ABRE 1 1 3 0 4 2 3 3 1 P$LEGB 6 4 8 2 1 2 2 2 7 P$NACF 2 2 2 1 1 0 3 3 2 P$OCSE 3 4 3 5 3 3 7 7 3 No of occurences p- p- p- p- p- BnPEF- BnPEF- p- BnRTI-4 BnMTFL BnGSTF var1 var2 BnLSP TF- (SEQ- (SEQ-ID (SEQ-ID (SEQ-ID (SEQ-ID (SEQ- Family ID 103) 111) 119) 6) 9) ID 125) O$VTBP 9 10 13 28 4 12 P$AHBP 22 9 9 18 5 14 P$DOFF 8 6 4 10 8 5 P$GTBX 8 10 5 22 5 10 P$IBOX 3 1 1 8 2 3 P$L1BX 10 2 1 10 3 9 P$LREM 6 2 2 8 4 5 P$MYBL 10 9 15 18 5 6 P$MYBS 13 3 4 5 3 5 P$NCS1 1 3 2 1 1 2 P$CCAF 4 3 3 9 2 0 P$GBOX 12 1 5 2 0 7 P$IDDF 1 3 3 3 1 0 P$MADS 12 6 3 8 5 6 P$MYCL 3 1 6 4 1 1 P$OPAQ 12 0 5 4 2 7 P$SPF1 3 0 2 7 1 2 P$SUCB 1 2 1 7 2 3 P$WBXF 1 2 1 3 1 5 O$INRE 2 4 2 7 2 2 O$PTBP 4 3 8 7 2 0 P$ABRE 3 1 3 1 0 4 P$LEGB 3 0 1 2 0 2 P$NACF 3 1 0 1 1 1 P$OCSE 7 0 4 4 0 3

All references cited in this specification are herewith incorporated by reference with respect to their entire disclosure content and the disclosure content specifically mentioned in this specification.

The invention will now be illustrated by the following Examples which are not intended, whatsoever, to limit the scope of this application.

Example 1 General Cloning Methods

General Cloning Methods including enzymatic digestion by restriction enzymes, agarous gel electrophoresys, purification of DNA fragments, transfer of nucleic acids to nitrocellulose on nylon membranes, maligation of DNA fragments, transformation of E. coli bacteria as well as culture of bacteria and sequence analysis of recombinant DNA have been carried out as described in Sambrook et al. (1989, Cold Spring Harbour Laboratory Press. ISBN 0-87969-309-6).

Example 2 Cloning of Promotor Elements from Brassica napus

For the analysis of seed specific expressed genes in Brassica napus, different tissues of various developmental stages (table 10) have been investigated.

TABLE 10 tissue types used for transcript analysis Specific developmental stages are given using the Biologische Bundesanstalt, Bundessortenamt, and Chemical Industry (BBCH) code (Meier, 1997) Sample Developmental Nr. Tissue stage type sample 1 immature embryos walking stick purpose prio 1 2 immature embryos fully developped purpose prio 1 (approx 20 days) 3 immature embryos fully developped purpose prio 1 pre dessic. green 4 immature embryos fully developped. control/purpose start dessic Yellow prio 2 5 embryo sac complete heart control/purpose prio 2 6 immature seed coat and weight mix control/purpose endosperm sample 1-3 prio 2 7 flower buds BBCH 57 control/purpose prio 2 8 anthers and stigma BBCH 68 control/purpose prio 2 9 flowers (exclude anthers BBCH 67 control and stigma) 10 leaves BBCH 32 control 11 leaves BBCH 75 control 12 leaves BBCH 80, control senescing (yellow) 13 stem BBCH 32 control 14 stem BBCH 75 control 15 stem BBCH 80, control senescing (yellow) 16 seedling: hypocotyl and BBCH 12 control cotyledons 17 seedling: roots (sand) BBCH 12 control 18 empty siliques weight mix control sample 1-3

To this end, Brassica napus cv. Kumily plants were raised under standard conditions (Moloney et al. 1992, Plant Cell Reports 8: 238-242). The tissues where harvested at the indicated developmental stages (table 10) and used for the preparation of RNA (RNAeasy, Qiagen) according to the manufactures manual. To identify in further experiments seed specifically expressed mRNA transcripts, three pools were created by mixing the RNA. A seed specific pool P-S was created from sample 1, 2, 3 (see table 10). A control pool P-C1 was created from sample 9, 10, 11, 12, 13, 14, 15, 16, 17, 18. A third pool P-C2 consisting of sample 4, 5, 6 7, 8 was used as a more stringent control: transcripts not expressed in control pool P-C2 are solely expressed in early seed development and in no other tissue/developmental stage. Using the amplified fragment length polymorphism (AFLP) method, known to a person skilled in the art, 384 primer combinations were used to identify 96 candidate transcript fragments being present solely in pool P-S and/or also weakly in pool P-C2 but absent on pool P-C1. A selection of 42 primer combinations that identified the 55 most promising transcript fragments were used to analyse in detail all samples listed in table 10, resulting in 26 fragment with confirmed expression solely in the developing embryo samples 1, 2, 3 and 4. Sequencing of those candidate fragments resulted in 20 unique sequences. Basic Local Alignment Search Tool (BLAST) was used to identify the putative full length transcripts corresponding to the identified transcript fragments. Surprisingly, besides genes known in the art for seed specific expression, e.g. napin and 3-ketoacyl-CoA synthase, a number of fragments listed in table 11 showed no homology to any known Brassica sequence, or to genes which were not known to be seed specifically expressed and/or to genes with unknown sequence upstream of the known mRNA sequence.

TABLE 11 Candidate fragments and homologue sequences identified using BLAST. fragment SEQ- Brassica Arabidopsis Candidate length ID homologue homologue BnSCP 157 bp 58 At5g36100 (unknown protein) BnGRPL 174 bp 4 At3g10185 (similar to gibberelin responsive protein) BnCRU4 127 bp 59 BnCRU4rn (Cruciferin) BnMYR 101 bp 60 BnMYRmc (Myrosinase) BnSETL 120 bp 26 At1g03270 (unknown protein) BnSCT2 172 bp 19 BnSCT2 (Sinapoyl-cholin transferase 2) BnMDP 273 bp 61 At3g20370 (unknown, similar BnRTI-4 237 bp 62 BnRTI-4 (Trypsin Inhibitor family) BnMTFL 144 bp 63 At2g21650 (put. Myb factor transcr. Factor) BnGSTF 186 bp 64 At3g62760 (AtGSTF13) BnSRP 131 bp 65 BnPEF 121 bp 12 Pectinesterase At5g47500 family protein (pectinesterase family protein) BnWSP  83 bp 66 BnLSP 164 bp 67 At5g62200 (weak similarity to embryo- specific protein 3)

From leaf material of Brassica napus cv. Kumily, genomic DNA has been isolated using the DNAeasy kit (Qiagen) according to the manufacturer's manual. Culture conditions for the Brassica napus cv. Kumily were as discussed above. Using this genomic DNA, as template, multiple rounds of thermal asymmetric interlaced polymerase chain reaction (TAIL-PCR)—a method known to a person skilled in the art—was performed to isolate sequences 5′ upstream and 3′ downstream of the 20 identified expressed fragments. The amplified products were either sequences directly, or supcloned into the pGEM-T (Promega) vector prior sequencing. Subcloning was required in some cases, as Brassica napus is amphidiploid, that is, Brassica napus contains 10 chromosomes common with Brassica rapa (A genome) and 9 chromosomes common with Brassica oleracea (C genome). Subcloning of PCR products containing a mixture of two sequences amplified from both genomes allows to separate these two sequences. Sequencing was done by standard techniques (laser fluorescent DNA-sequenceing, ABI according to the method of Sanger et al. 1977 Proc. Natl. Acad. Sci. USA 74, 5463-5467).

For candidate expressed sequences where no Brassica gene was known to the art, the open reading frame was identified with help of software prediction and using alignments with homologues genes from Arabidopsis. The following Brassica napus open reading frames have been identified (Table 12):

TABLE 12 Open reading frames (ORF) identified in Brassica napus cDNA sequences. Brassica napus Sequence ORF in bp SEQ ID NO: BnGRPL 360 5 BnPEF 1083 13 BnSCT1 1401 20 BnSCT2 1401 21 BnSETL1 1512 27 BnSETL2 1512 28 BnSCP 773 72 BnCRU4 1107 80 BnMYR 1644 88 BnMDP 1152 98 BnRTI-4 300 106 BnMTFL 303 114 BnGSTF 660 121 BnLSP 574 133

For the expressed sequence SEQ-ID 126, no open reading frame could be identified.

The following Brassica napus sequences upstream of the expressed sequence SEQ-ID 126 or the identified expressed open reading frames (ORF) have been obtained (Table 13).

TABLE 13 Genomic 5′ upstream sequences from the Brassica napus cDNA sequences. Brassica napus genomic 5′ Sequence sequence in bp SEQ ID NO: p-BnGRPL 1790 1 p-BnPEF-var1 2027 6 p-BnPEF-var2 636 9 p-BnSCT2-var1 1019 14 p-BnSCT2-var2 996 16 p-BnSETL-var1 1490 22 p-BnSETL-var2 2010 25 p-BnSCP 2052 70 p-BnCRU4 1951 77 p-BnMYR 1360 85 p-BnMDP 1428 95 p-BnRTI-4 1820 103 p-BnMTFL 1335 111 p-BnGSTF 1565 119 p-BnSRP 2447 124 p-BnLSP 1593 121

The analysis of the 5′ upstream sequences using Genomatix software Gems-Launcher showed that the sequences comprised promoter elements. This was confirmed by the presence of a TATA-Box which is required for transcription by RNA-polymerases. Also in the isolated fragments elements specific for seed-transcription factors (e.g. Prolaminbox, legumin box, RITA etc.) were found.

The following Brassica napus sequences downstream of the identified expressed open reading frames (ORF) have been obtained (Table 14).

TABLE 14 Genomic 3′ downstream sequences from the Brassica napus cDNA sequences. Brassica napus genomic 3′ Sequence sequence in bp SEQ ID NO: t-BnGRPL 581 2 t-BnPEF-var1 477 7 t-BnPEF-var2 538 10 t-BnSCT2-var1 573 15 t-BnSCT2-var2 576 17 t-BnSETL-var1 614 23 t-BnSCP 587 71 t-BnCRU4 514 78 t-BnMYR 652 86 t-BnMDP 483 96 t-BnRTI-4 572 104 t-BnMTFL 521 112 t-BnSRP 865 125

Example 3 Production of Test Constructs for Demonstrating Promoter Activity

For the testing of the promoter elements in a first step promoter terminator cassettes were generated. To this end, fusion PCRs have been used wherein via two PCR steps promoter elements were linked with terminator elements. At the same time, a multiple cloning site was introduced in between the promoter and terminator elements. The primers used to generate cassettes using corresponding native Brassica terminators are shown in Table 15, Table 16 list the generated cassettes using the OCS terminator.

TABLE 15 Primer pairs used for the generation of promoter-multiple cloning site- terminated-cassettes via Fusion-PCR using native Brassica terminator sequences. Brassica napus Promoter/Terminator Primer pair 1. PCR Primer pair 1. PCR cassette Promoter Terminator Primer pair 2. PCR p-BnGRP_MCS_t- Forw: atacccggga- Forw: ccatggctc- Forw: atacccgggatac BnGRP tacctgcaggt- gatcgttaattaaag- ctgcaggttaggccggc- taggccggccacaa- catgcgaggcctataatt- cacaaaacgaaatcat- aacgaaatcatca- gattgtttctcctcttagtttg- caaatcgtg (SEQ ID aatcgtg (SEQ ID taaataatctatc (SEQ NO: 29) NO: 29) ID NO: 31) Rev: ‘taagcggccg- Rev: caatcaatta- Rev: taagcggccg- caatcggaccga- taggcctcgcatgctt- caatcggaccga- taccggtaggcgccgat- taattaacgatcgagc- taccggtaggcgccgat- tatctacctaacctaaca- catggttttagagagccg- tatctacctaacctaaca- aacaaaag (SEQ ID gaattattg (SEQ ID aacaaaag (SEQ ID NO: 32) NO: 30) NO: 32) p-BnPEF- Forw: Forw: Forw: var1_MCS_t- atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca BnPEF-var1 ggttaggccggccaaattt aaagcatgcgaggcctat ggttaggccggccaaattt attaacccatctatttgttca aattgattgtaaggaatca attaacccatctatttgttca c (SEQ ID NO: 33) acttcaaatgcttttc c (SEQ ID NO: 33) Rev: (SEQ ID NO: 35) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatgggacgagaaa accgataccggtaggcg ccgacgttaagcgagagt gaaaatggtcggag ccgacgttaagcgagagt ggatag (SEQ ID (SEQ ID NO: 34) ggatag (SEQ ID NO: 36) NO: 36) p-BnPEF- Forw: Forw: Forw: var2_MCS_t- atacccgggatacctgca Ccatggctcgatcgttaat atacccgggatacctgca BnPEF-var2 ggttaggccggccagaat taaagcatgcgaggccta ggttaggccggccagaat tacgtttgagttcaaattca taattgattgtaggaatca tacgtttgagttcaaattca g (SEQ ID NO: 37) acttcaaatgctttt (SEQ g (SEQ ID NO: 37) Rev: ID NO: 39) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatgggacgagaaa accgataccggtaggcg ccagaggtgaggaggag taaatggtcgaag (SEQ ccagaggtgaggaggag ttgcac (SEQ ID ID NO: 38) ttgcac (SEQ ID NO: 40) NO: 40) p-BnSCT2- Forw: Forw: Forw: var1_MCS_t- atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca BnSCT2-var1 ggttaggccggccagatg aaagcatgcgaggcctat ggttaggccggccagatg caaaaacgtatagtcaca aattgattgAgttcctcact caaaaacgtatagtcaca c (SEQ ID NO: 41) cacttctctc (SEQ ID c (SEQ ID NO: 41) Rev: NO: 43) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggtttctctgcttct accgataccggtaggcg ccgccttatatggattttgtg tggtgtcac (SEQ ID ccgccttatatggattttgtg ttactgacc (SEQ ID NO: 42) ttactgacc (SEQ ID NO: 44) NO: 44) p-BnSCT2- Forw: Forw: Forw: var2_MCS_t- atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca BnSCT2-var2 ggttaggccggccagatg aaagcatgcgaggcctat ggttaggccggccagatg caaaaacgtatagtcaca aattgattgAgttcctcact caaaaacgtatagtcaca c (SEQ ID NO: 45) cacttctctc (SEQ ID c (SEQ ID NO: 45) Rev: NO: 47) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggtttctctgcttct accgataccggtaggcg ccgccttatatggattttgtg tggtgtcac (SEQ ID ccgccttatatggattttgtg ttactgacc (SEQ ID NO: 46) ttactgacc (SEQ ID NO: 48) NO: 48) p-BnSETL- Forw: Forw: Forw: var1_MCS_t- atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca BnSETL-var1 ggttaggccggccagtag aaagcatgcgaggcctat ggttaggccggccagtag aagttattagcaacttgtac aattgattgtacatactatat aagttattagcaacttgtac acac (SEQ ID tttttgtttaccttgtg (SEQ acac (SEQ ID NO: 49) ID NO: 51) NO: 49) Rev: Rev: Rev: caatcaattataggcctcg taagcggccgcaatcgg taagcggccgcaatcgg catgctttaattaacgatcg accgataccggtaggcg accgataccggtaggcg agccatggtttgaccccttc cccaaacacggctcaga cccaaacacggctcaga ttgttcttc (SEQ ID gaagc (SEQ ID gaagc (SEQ ID NO: 50) NO: 52) NO: 52) p-BnSETL- Forw: Forw: Forw: var2_MCS_t- atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca BnSETL-var1 ggttaggccggccatcgg aaagcatgcgaggcctat ggttaggccggccatcgg ctacaaatccaactgg aattgattgtacatactatat ctacaaatccaactgg (SEQ ID NO: 53) tttttgtttaccttgtg (SEQ (SEQ ID NO: 53) Rev:- ID NO: 51) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggtttgtcgttttcc accgataccggtaggcg cccaaacacggctcaga tcagcttc (SEQ ID cccaaacacggctcaga gaagc (SEQ ID NO: 54) gaagc (SEQ ID NO: 52) NO: 52) p-BnSCP_MCS_t- Forw: Forw: Forw: BnSCP atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca ggttaggccggccaatca aaagcatgcgaggcctat ggttaggccggccaatca taagttgtatcagttcatc aattgattgggagaaaat taagttgtatcagttcatc (SEQ-ID No: 73) atgggagaagatggaa (SEQ-ID No: 73) Rev: (SEQ-ID No: 75) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggtgtttgactcat accgataccggtaggcg cctatagacctgccaaatc actggtggta (SEQ-ID cctatagacctgccaaatc aaaccaac (SEQ-ID No: 74) aaaccaac (SEQ-ID No: 76) No: 76) p- Forw: Forw: Forw: BnCRU4_MCS_t- atacccgggatacc- ccatggctcgatcgttaat- atacccgggatacc- BnCRU4 tgcaggttagg- taaagcatgcgagg- tgcaggttagg- ccggccaatgtacatg- cctataattgattgagcat- ccggccaatgtacatg- gatgcgtatagatg gagttagtgatgtaa- gatgcgtatagatg (SEQ-ID No: 81) cagcg (SEQ-ID (SEQ-ID No: 81) Rev: No: 83) Rev: caatcaattatagg- Rev: taagcggccgcaatc- cctcgcatgctttaat- taagcggccgcaatc- ggaccgatacc- taacgatcgagccatg- ggaccgatacc- ggtaggcgccggaaga- gtagagcatg- ggtaggcgccggaaga- gatggaagcttaca- tccgtgaacgacg gatggaagcttaca- gaatg (SEQ-ID (SEQ-ID No: 82) gaatg (SEQ-ID No: 84) No: 84) p-BnMYR_MCS_t- Forw: Forw: Forw: BnMYR atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca ggttaggccggccacata aaagcatgcgaggcctat ggttaggccggccacata aaaatatgttgagaaaat aattgattgaacactttatc aaaatatgttgagaaaat atc (SEQ-ID No: 89) cacatcaagatcgc atc (SEQ-ID No: 89) Rev: (SEQ-ID No: 91) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggggttaatgtgt accgataccggtaggcg ccgatgtgtgctcgtaattg agatttgtatatatg ccgatgtgtgctcgtaattg cactttt (SEQ-ID (SEQ-ID No: 90) cactttt (SEQ-ID No: 92) No: 92) p-BnMDP_MCS_t- Forw: Forw: Forw: BnMDP atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca ggttaggccggccatgcg aaagcatgcgaggcctat ggttaggccggccatgcg agagagaagaaatgaa aattgattgatttctctaact agagagaagaaatgaa ataag (SEQ-ID aagaaactttgtag ataag (SEQ-ID No: 99) (SEQ-ID No: 101) No: 99) Rev: Rev: Rev: caatcaattataggcctcg taagcggccgcaatcgg taagcggccgcaatcgg catgctttaattaacgatcg accgataccggtaggcg accgataccggtaggcg agccatggcttttgagattg cccgataaagatggtcta cccgataaagatggtcta tatatatgaatg (SEQ- atgtccatc (SEQ-ID atgtccatc (SEQ-ID ID No: 100) No: 102) No: 102) p-BnRTI- Forw: Forw: Forw: 4_MCS_t-BnRTI-4 atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca ggttaggccggccacttg aaagcatgcgaggcctat ggttaggccggccacttg cgccgaagatatatccga aattgattgatacttcacaa cgccgaagatatatccga c (SEQ-ID No: 107) ctttgcataagcc c (SEQ-ID No: 107) Rev: (SEQ-ID No: 109) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggcttctctctata accgataccggtaggcg ccaccaaatccgaaacc tatctcttac (SEQ-ID ccaccaaatccgaaacc gaatccgaac (SEQ- No: 108) gaatccgaac (SEQ- ID No: 110) ID No: 110) p- Forw: Forw: Forw: BnMTFL_MCS_t- atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca BnMTFL ggttaggccggccatattc aaagcatgcgaggcctat ggttaggccggccatattc actacttatagagaacac aattgattgaacaagaag actacttatagagaacac (SEQ-ID No: 115) caacaaagcctaaactat (SEQ-ID No: 115) Rev: (SEQ-ID No: 117) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggtgtgaaaaag accgataccggtaggcg ccgatttgagttttcgtgaa tttggcgtatttc (SEQ- ccgatttgagttttcgtgaa gttcaga (SEQ-ID ID No: 116) gttcaga (SEQ-ID No: 118) No: 118) p-BnSRP_MCS_t- Forw: Forw: Forw: BnSRP atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca ggttaggccggccaaac aaagcatgcgaggcctat ggttaggccggccaaac ggcaattgattctcgccct aattgattgtttacatattgg ggcaattgattctcgccct g (SEQ-ID No: 127) cccaagaggcataat g (SEQ-ID No: 127) Rev: (SEQ-ID No: 129) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggagagaggtta accgataccggtaggcg ccgttaatgaaaacggta ttgaaaccacgt (SEQ- ccgttaatgaaaacggta ccgaatttc (SEQ-ID ID No: 128) ccgaatttc (SEQ-ID No: 130) No: 130)

TABLE 16 Primer pairs used for the generation of promoter-multiple cloning site- terminated-cassettes via Fusion-PCR using the OCS terminator sequence. p-BnGRP_MCS_t- Forw: atacccggga- Forw: ccatg- Forw: atacccggga- OCS tacctgcaggttagg- gctcgatcgttaat- tacctgcaggttagg- ccggccacaaaac- taaagcatgcgagg- ccggccacaaaac- gaaatcatcaaatcgtg cctataattgatt- gaaatcatcaaatcgtg (SEQ ID NO: 29) gctgctttaatgagatatg- (SEQ ID NO: 29) Rev: caatcaatta- cgagacg (SEQ ID Rev: taagc- taggcctcgcatg- NO: 55) ggccgcaatcggacc- ctttaattaacgatc- Rev: taagc- gataccggtagg- gagccatggttttaga- ggccgcaatcggacc- cgccacaatcag- gagccggaattattg gataccggtagg- taaattgaacggagaa- (SEQ ID NO: 30) cgccacaatcag- tattattc (SEQ ID taaattgaacggagaa- NO: 56) tattattc (SEQ ID NO: 56) p-BnPEF- Forw: atacccggga- Forw: ccatg- Forw: atacccggga- var1_MCS_t-OCS tacctgcaggttagg- gctcgatcgttaat- tacctgcaggttagg- ccggccaaatttat- taaagcatgcgagg- ccggccaaatttat- taacccatctatttgttcac cctataattgatt- taacccatctatttgttcac (SEQ ID NO: 33) gctgctttaatgagatatg- (SEQ ID NO: 33) Rev: caatcaatta- cgagacg (SEQ ID Rev: taagc- taggcctcgcatg- NO: 55) ggccgcaatcggacc- ctttaattaacgatc- Rev: taagc- gataccggtagg- gagccatgggacga- ggccgcaatcggacc- cgccacaatcag- gaaagaaaatg- gataccggtagg- taaattgaacggagaa- gtcggag (SEQ ID cgccacaatcag- tattattc (SEQ ID NO: 34) taaattgaacggagaa- NO: 56) tattattc (SEQ ID NO: 56) p-BnPEF- Forw: atacccggga- Forw: ccatg- Forw: atacccggga- var2_MCS_t-OCS tacctgcaggttagg- gctcgatcgttaat- tacctgcaggtagg- ccggccagaattacg- taaagcatgcgagg- ccggccagaattacg- tttgagttcaaattcag cctataattgatt- tttgagttcaaattcag (SEQ ID NO: 37) gctgctttaatgagatatg- (SEQ ID NO: 37) Rev: caatcaatta- cgagacg (SEQ ID Rev: taagc- taggcctcgcatg- NO: 55) ggccgcaatcggacc- ctttaattaacgatc- Rev: taagc- gataccggtagg- gagccatgggacga- ggccgcaatcggacc- cgccacaatcag- gaaataaatggtcgaag gataccggtagg- taaattgaacggagaa- (SEQ ID NO: 38) cgccacaatcag- tattattc (SEQ ID taaattgaacggagaa- NO: 56) tattattc (SEQ ID NO: 56) p-BnSCT2- Forw: Forw: Forw: var1_MCS_t-OCS atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca ggttaggccggccagatg aaagcatgcgaggcctat ggttaggccggccagatg caaaaacgtatagtcaca aattgattgctgctttaatga caaaaacgtatagtcaca c (SEQ ID NO: 41) gatatgcgagacg c (SEQ ID NO: 41) Rev: (SEQ ID NO: 55) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggtttctctgcttct accgataccggtaggcg ccacaatcagtaaattga tggtgtcac (SEQ ID ccacaatcagtaaattga acggagaatattattc NO: 42) acggagaatattattc (SEQ ID NO: 56) (SEQ ID NO: 56) p-BnSCT2- Forw: Forw: Forw: var2_MCS_t-OCS atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca ggttaggccggccagatg aaagcatgcgaggcctat ggttaggccggccagatg caaaaacgtatagtcaca aattgattgctgctttaatga caaaaacgtatagtcaca c (SEQ ID NO: 45) gatatgcgagacg c (SEQ ID NO: 45) Rev: (SEQ ID NO: 55) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggtttctctgcttct accgataccggtaggcg ccacaatcagtaaattga tggtgtcac (SEQ ID ccacaatcagtaaattga acggagaatattattc NO: 46) acggagaatattattc (SEQ ID NO: 56) (SEQ ID NO: 56) p-BnSETL- Forw: Forw: Forw: var1_MCS_t-OCS atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca ggttaggccggccagtag aaagcatgcgaggcctat ggttaggccggccagtag aagttattagcaacttgtac aattgattgctgctttaatga aagttattagcaacttgtac acac (SEQ ID gatatgcgagacg acac (SEQ ID NO: 49) (SEQ ID NO: 55) NO: 49) Rev: Rev: Rev: caatcaattataggcctcg taagcggccgcaatcgg taagcggccgcaatcgg catgctttaattaacgatcg accgataccggtaggcg accgataccggtaggcg agccatggtttgaccccttc ccacaatcagtaaattga ccacaatcagtaaattga ttgttcttc (SEQ ID acggagaatattattc acggagaatattattc NO: 50) (SEQ ID NO: 56) (SEQ ID NO: 56) p-BnSETL- Forw: Forw: Forw: var2_MCS_t-OCS atacccgggatacctgca ccatggctcgatcgttaatt atacccgggatacctgca ggttaggccggccatcgg aaagcatgcgaggcctat ggttaggccggccatcgg ctacaaatccaactgg aattgattgctgctttaatga ctacaaatccaactgg (SEQ ID NO: 53) gatatgcgagacg (SEQ ID NO: 53) Rev:- (SEQ ID NO: 55) Rev: caatcaattataggcctcg Rev: taagcggccgcaatcgg catgctttaattaacgatcg taagcggccgcaatcgg accgataccggtaggcg agccatggtttgtcgttttcc accgataccggtaggcg ccacaatcagtaaattga tcagcttc (SEQ ID ccacaatcagtaaattga acggagaatattattc NO: 54) acggagaatattattc (SEQ ID NO: 56) (SEQ ID NO: 56) p-BnSCP_MCS_t- Forw: Forw: Forw: OCS atacccgggatacctg- ccatggctcgatcgttaat- atacccgggatacctg- caggttaggccggc- taaagcatgcgagcc- caggttaggccggc- caatcataagttgtat- tataattgattgctgctt- caatcataagttgtat- cagttcatc (SEQ ID taatgagatatgcga- cagttcatc (SEQ-ID No: 73) gacg (SEQ-ID No: 73) Rev: No: 55) Rev: caatcaatta- Rev: taagcggccgcaatcg- taggcctcgcatgctt- taagcggccgcaatcg- gaccgataccgg- taattaacgatcgagc- gaccgataccgg- taggcgccacaatcag- catggtgtttgactca- taggcgccacaatcag- taaattgaacggagaa- tactggtggta (SEQ-ID taaattgaacggagaa- tattattc (SEQ-ID No: 74) tattattc (SEQ-ID No: 56) No: 56) p- Forw: Forw: Forw: BnCRU4_MCS_t- atacccgggatacc- ccatggctcgatcgttaat- atacccgggatacc- OCS tgcaggttagg- taaagcatgcgagg- tgcaggttagg- ccggccaatgtacatg- cctataattgatt- ccggccaatgtacatg- gatgcgtatagatg cctataattgatt- gatgcgtatagatg (SEQ-ID No: 81) gctgctttaatgagatatg- (SEQ-ID No: 81) Rev: cgagacg (SEQ-ID Rev: caatcaattatagg- No: 55) taagcggccgcaatc- cctcgcatgctttaat- Rev: ggaccgatacc- taacgatcgagccatg- taagccggccgcaatc- ggtaggcgccacaat- gtagagcatg- ggaccgatacc- cagtaaattgaacgga- tccgtgaacgacg ggtaggcgccacaat- gaatattattc (SEQ-ID (SEQ-ID No: 82) cagtaaattgaacgga- No: 56) gaatattattc (SEQ-ID No: 56) p-BnMYR_MCS_t- Forw: Forw: Forw: OCS atacccgggatacc- ccatggctcgatcgttaat- atacccgggatacc- tgcaggttagg- taaagcatgcgagg- tgcaggttagg- ccggccacataaaaa- cctataattgatt- ccggccacataaaaa- tatgttgagaaaatatc gctgctttaatgagatatg- tatgttgagaaaatatc (SEQ-ID No: 89) cgagacg (SEQ-ID (SEQ-ID No: 89) Rev: No: 55) Rev: caatcaattatagg- Rev: taagcggccgcaatc- cctcgcatgctttaat- taagcggccgcaatc- ggaccgatacc- taacgatcgagccatg- ggaccgatacc- ggtaggcgccacaat- gggttaatgtgtagatt- ggtaggcgccacaat- cagtaaattgaacgga- tgtatatatg (SEQ-ID cagtaaattgaacgga- gaatattattc (SEQ-ID No: 90) gaatattattc (SEQ-ID No: 56) No: 56) p-BnMDP_MCS_t- Forw: Forw: Forw: OCS atacccgggatacctg- ccatggctcgatcgttaat- atacccgggatacctg- caggttaggccggc- taaagcatgcgaggcc- caggttaggccggc- catgcgagagagaaga- tataattgattgctgctt- catgcgagagagaaga- aatgaaataag (SEQ- taatgagatatgcga- aatgaaataag (SEQ- ID No: 99) gacg (SEQ-ID ID No: 99) Rev: No: 55) Rev: caatcaatta- Rev: taagcggccgcaatcg- taggcctcgcatgctt- taagcggccgcaatcg- gaccgataccgg- taattaacgatcgagc- gaccgataccgg- taggcgccacaatcag- catggcttttgagattgta- taggcgccacaatcag- taaattgaacggagaa- tatatgaatg (SEQ-ID taaattgaacggagaa- tattattc (SEQ-ID No: 100) tattattc (SEQ-ID No: 56) No: 56) p-BnRTI- Forw: Forw: Forw: 4_MCS_t-OCS atacccgggatacc- ccatggctcgatcgttaat- atacccgggatacc- tgcaggttagg- taaagcatgcgagg- tgcaggttagg- ccggccact- cctataattgatt- ccggccact- tgcgccgaagatatatc- gctgctttaatgagatatg- tgcgccgaagatatatc- cgac (SEQ-ID cgagacg (SEQ-ID cgac (SEQ-ID No: 107) No: 55) No: 107) Rev: Rev: Rev: caatcaattatagg- taagcggccgcaatc- taagcggccgcaatc- cctcgcatgctttaat- ggaccgatacc- ggaccgatacc- taacgatcgagccatg- ggtaggcgccacaat- ggtaggcgccacaat- gcttctctctatatatc- cagtaaattgaacgga- cagtaaattgaacgga- tcttac (SEQ-ID gaatattattc (SEQ-ID gaatattattc (SEQ-ID No: 108) No: 56) No: 56) p Forw: Forw: Forw: BnMTFL_MCS_t- atacccgggatacc- ccatggctcgatcgttaat- atacccgggatacc- OCS tgcaggttagg- taaagcatgcgagg- tgcaggttagg- ccggccatattcactact- cctataattgatt- ccggccatattcactact- tatagagaacac (SEQ- gctgctttaatgagatatg- tatagagaacac (SEQ- ID No: 115) cgagacg (SEQ-ID ID No: 115) Rev: No: 55) Rev: caatcaattatagg- Rev: taagcggccgcaatc- cctcgcatgctttaat- taagcggccgcaatc- ggaccgatacc- taacgatcgagccatg- ggaccgatacc- ggtaggcgccacaat- gtgtgaaaaagt- ggtaggcgccacaat- cagtaaattgaacgga- ttggcgtatttc (SEQ-ID cagtaaattgaacgga- gaatattattc (SEQ-ID No: 116) gaatattattc (SEQ-ID No: 56) No: 56) p- Forw: Forw: Forw: BnGSTF_MCS_t- atacccgggatacc- ccatggctcgatcgttaat- atacccgggatacc- OCS tgcaggttagg- taaagcatgcgagg- tgcaggttagg- ccggccagagcattatg- cctataattgatt- ccggccagagcattatg- cttccaagcggac gctgctttaatgagatatg- cttccaagcggac (SEQ-ID No: 122) cgagacg (SEQ-ID (SEQ-ID No: 122) Rev: No: 55) Rev: caatcaattatagg- Rev: taagcggccgcaatc- cctcgcatgctttaat- taagcggccgcaatc- ggaccgatacc- taacgatcgagccatg- ggaccgatacc- ggtaggcgccacaat- gttcactctaact- ggtaggcgccacaat- cagtaaattgaacgga- tcgtaactcg (SEQ-ID cagtaaattgaacgga- gaatettattc (SEQ-ID No: 123) gaatattattc (SEQ-ID No: 56) No: 56) p-BnSRP_MCS_t- Forw: Forw: Forw: OCS atacccgggatacctg- ccatggctcgatcgttaat- atacccgggatacctg- caggttaggccggcca- taaagcatgcgaggcc- caggttaggccggcca- aacggcaatt- tataattgattgctgctt- aacggcaatt- gattctcgccctg (SEQ- taatgagatatgcga- gattctcgccctg (SEQ- ID No: 127) gacg (SEQ-ID ID No: 127) Rev: No: 55) Rev: caatcaatta- Rev: taagcggccgcaatcg- taggcctcgcatgctt- taagcggccgcaatcg- gaccgataccgg- taattaacgatcgagc- gaccgataccgg- taggcgccacaatcag- catggagagaggttatt- taggcgccacaatcag- taaattgaacggagaa- gaaaccacgt (SEQ- taaattgaacggagaa- tattattc (SEQ-ID ID No: 128) tattattc (SEQ-ID No: 56) No: 56) p-BnLSP_MCS_t- Forw: Forw: Forw: OCS atacccgggatacctg- ccatggctcgatcgttaat- atacccgggatacctg- caggttaggccggcca- taaagcatgcgaggcc- caggttaggccggcca- aatagtcaagtttatgaat- tataattgattgctgctt- aatagtcaagtttatgaat- cacag (SEQ-ID taatgagatatgcga- cacag (SEQ-ID No: 134) gacg (SEQ-ID No: 134) Rev: No: 55) Rev: caatcaatta- Rev: taagcggccgcaatcg- taggcctcgcatgctt- taagcggccgcaatcg- gaccgataccgg- taattaacgatcgagc- gaccgataccgg- taggcgccacaatcag- catggtcttgaacttctt- taggcgccacaatcag- taaattgaacggagaa- gacattact (SEQ-ID taaattgaacggagaa- tattattc (SEQ-ID No: 135) tattattc (SEQ-ID No: 56): No: 56) p-Napin_MCS_t- Forw: Forw: Forw: OCS atacccgggatacc- ccatggctcgatcgttaat- atacccgggatacc- tgcaggttagg- taaagcatgcgagg- tgcaggttagg- ccggccataaggat- cctataattgatt- ccggccataaggat- gacctacccattcttga gctgctttaatgagatatg- gacctacccattcttga (SEQ-ID No: 138) cgagacg (SEQ-ID (SEQ-ID No: 138) Rev: No: 55) Rev: caatcaattatagg- Rev: taagcggccgcaatc- cctcgcatgctttaat- taagcggccgcaatc- ggaccgatacc- taacgatcgagccatg- ggaccgatacc- ggtaggcgccacaat- gtgtttttaatcttgtttgtatt ggtaggcgccacaat- cagtaaattgaacgga- (SEQ-ID No: 139) cagtaaattgaacgga- gaatattattc (SEQ-ID gaatattattc (SEQ-ID No: 56) No: 56) p-LuPXR_MCS_t- Forw: Forw: Forw: OCS atacccgggatacc- ccatggctcgatcgttaat- atacccgggatacc- tgcaggttagg- taaagcatgcgagg- tgcaggttagg- ccggccacacgggcag- cctataattgatt- ccggccacacgggcag- gacatagggactact gctgctttaatgagatatg- gacatagggactact (SEQ-ID No: 142) cgagacg (SEQ-ID (SEQ-ID No: 142) Rev: No: 55) Rev: caatcaattatagg- Rev: taagcggccgcaatc- cctcgcatgctttaat- taagcggccgcaatc- ggaccgatacc- taacgatcgagccatg- ggaccgatacc- ggtaggcgccacaat- ggatttatgataaaaatg- ggtaggcgccacaat- cagtaaattgaacgga- tcggt (SEQ-ID cagtaaattgaacgga- gaatattattc (SEQ-ID No: 143) gaatattattc (SEQ-ID No: 56) No: 56)

The promoter-terminator cassettes were cloned into the pCR2.1 (Invitrogen) vector according to the manufacturer's manual and subsequently sequenced. In a further step, the delta 6 Desaturase Gene (SEQ ID NO: 68) was introduced via the NcoI, PacI restrictions site between the promoter and terminator sequence.

Using the Multisite Gateway System (Invitrogen), a multiple cloning site (SEQ ID 57) was introduced into each of the three pENTR vectors pENTR/A pENTR/B and pENTR/C via HindIII and KpnI restrictions sites. Into the first position of this MCS, the promotor-delta 6 Desaturase-terminator cassette was cloned via FseI and KasI. Similarily, the DsRed gene was introduced into pENTR/C between the Napin promotor and the OCS terminator. By performing a site specific recombination (LR-reaction), the created pENTR/B, pENTR/C and an empty pENTR/A vector were combined with the pSUN destination vector according to the manufacturers (Invitrogen) Multisite Gateway manual to generate the final binary vectors SEQ ID: 3 pSUN-p-GRPL_d6Des(Pir)_t-OCS, SEQ ID: 8 pSUN-p-PEF-var1_d6Des(Pir)_t-OCS, SEQ ID: 11 pSUN-p-PEF-var2_d6Des(Pir)_t-OCS, SEQ ID: 18 pSUN-p-SCT2-var2_d6Des(Pir)_t-OCS, SEQ ID: 24 pSUN-p-SETL-vad_t-OCS, SEQ ID: 79 pSUN-pBnCRU4_d6Des(Pir)_t-OCS, SEQ ID: 87 pSUN-pBnMYR_d6Des(Pir)_t-OCS, SEQ ID: 93 pSUN-pBnSETL-var2_d6Des(Pir)_t-OCS, SEQ ID: 94 pSUN-pBnSCT2-vad_d6Des(Pir)_t-OCS, SEQ ID: 97 pSUN-pBnMDP_d6Des(Pir)_t-OCS, SEQ ID: 105 pSUN-pBnRTI-4_d6Des(Pir)_t-OCS, SEQ ID: 113 pSUN-pBnMTFL_d6Des(Pir)_t-OCS, SEQ ID: 120 pSUN-pBnGSTF_d6Des(Pir)_t-OCS, SEQ ID: 132 pSUN-pBnLSP_d6Des(Pir)_t-OCS. Similarly, the two binary vectors SEQ ID: 137 pSUN-pNapin_d6Des(Pir)_t-OCS and SEQ ID: 141 pSUN-pLuPXR_d6Des(Pir)_t-OCS were cloned as a positive control, that is, these two vectors are known to be capable to drive seed specific expression of PUFA genes, e.g. the delta-6-desaturase SEQ ID NO: 68.

The resulting vectors were subsequently used for the production of transgenic plants. The promoter activity in the transgenic plant seeds was measured based on the expression of delta 6 Desaturase and an observed modification in the lipid pattern of the seeds as described in example 5.

Example 4 Production of Transgenic Plants

a) Generation of Transgenic Rape Seed Plants (Amended Protocol According to Moloney et al. 1992, Plant Cell Reports, 8:238-242)

For the generation of transgenic rapeseed plants, the binary vectors were transformed into Agrobacterium tumefaciens C58C1:pGV2260 (Deblaere et al. 1984, Nucl. Acids. Res. 13: 4777-4788). For the transformation of rapeseed plants (cv. Kumily,) a 1:50 dilution of an overnight culture of positive transformed acrobacteria colonies grown in Murashige-Skoog Medium (Murashige and Skoog 1962 Physiol. Plant. 15, 473) supplemented by 3% saccharose (3MS-Medium) was used. Petiols or Hypocotyledones of sterial rapeseed plants were incubated in a petri dish with a 1:50 acrobacterial dilusion for 5-10 minutes. This was followed by a tree day co-incubation in darkness at 25° C. on 3MS-Medium with 0.8% bacto-Agar. After three days the culture was put on to 16 hours light/8 hours darkness weekly on MS-medium containing 500 mg/l Claforan (Cefotaxime-Natrium), 100 nM Imazetapyr, 20 mikroM Benzylaminopurin (BAP) and 1.6 g/l Glucose. Growing sprouts were transferred to MS-Medium containing 2% saccharose, 250 mg/l Claforan and 0.8% Bacto-Agar. Even after three weeks no root formation was observed, a growth hormone 2-Indolbutyl acid was added to the medium for enhancing root formation.

Regenerated sprouts have been obtained on 2MS-Medium with Imazetapyr and Claforan and were transferred to the green house for sprouting. After flowering, the mature seeds were harvested and analysed for expression of the Desaturase gene via lipid analysis as described in Qui et al. 2001, J. Biol. Chem. 276, 31561-31566.

b) Production of Transgenic Flax Plants

The production of transgenic flax plants can be carried out according to the method of Bell et al., 1999, In Vitro Cell. Dev. Biol. Plant 35(6):456-465 using particle bombardment. Acrobacterial transformation could be carried out according to Mlynarova et al. (1994), Plant Cell Report 13: 282-285.

Example 5 Lipid Extraction

Lipids can be extracted as described in the standard literature including Ullman, Encyclopedia of Industrial Chemistry, Bd. A2, S. 89-90 and S. 443-613, VCH: Weinheim (1985); Fallon, A., et al., (1987) “Applications of HPLC in Biochemistry” in: Laboratory Techniques in Biochemistry and Molecular Biology, Bd. 17; Rehm et al. (1993) Biotechnology, Bd. 3, Kapitel III: “Product recovery and purification”, S. 469-714, VCH: Weinheim; Better, P. A., et al. (1988) Bioseparations: downstream processing for Biotechnology, John Wiley and Sons; Kennedy, J. F., und Cabral, J. M. S. (1992) Recovery processes for biological Materials, John Wiley and Sons; Shaeiwitz, J. A., und Henry, J. D. (1988) Biochemical Separations, in: Ullmann's Encyclopedia of Industrial Chemistry, Bd. B3; Kapitel 11, S. 1-27, VCH: Weinheim; und Dechow, F. J. (1989) Separation and purification techniques in biotechnology, Noyes Publications.

Alternatively, extraction will be carried out as described in Cahoon et al. (1999) Proc. Natl. Acad. Sci. USA 96 (22):12935-12940, und Browse et al. (1986) Analytic Biochemistry 152:141-145. Quantitative and qualitative analysis of lipids or fatty acids are described in Christie, William W., Advances in Lipid Methodology, Ayr/Scotland: Oily Press (Oily Press Lipid Library; 2); Christie, William W., Gas Chromatography and Lipids. A Practical Guide—Ayr, Scotland: Oily Press, 1989, Repr. 1992, IX, 307 S. (Oily Press Lipid Library; 1); “Progress in Lipid Research, Oxford: Pergamon Press, 1 (1952)-16 (1977) u.d.T.: Progress in the Chemistry of Fats and Other Lipids CODEN.

Based on the analysed lipids, the expression of the Desaturase can be determined since the lipid pattern of successfully transformed plant seeds will differ from the pattern of control plant seeds. Seed specific expression of a deta-6-desaturase would result in formation of 18:3n-6 (GLA) and/or 18:4n-3 (SDA), depending on whether the delta 6 desaturase uses 18:2n-6 (LA) and/or 18:3n-3 (ALA) as substrate. Surprisingly, not only the two control promotors Napin and LuPXR harbored by the vectors SEQ ID: 137 pSUN-pNapin_d6Des(Pir)_t-OCS and SEQ ID: 141 pSUN-pLuPXR_d6Des(Pir)_t-OCS were capable to drive seed specific expression of the delta-6-desaturase as indicated by the formation of GLA, but also the promotors of the present invention (FIG. 2). Interestingly, the promotors influenced the ratio of the omega-3 fatty acid SDA to the omega-6 fatty acid GLA. 

The invention claimed is:
 1. An expression cassette comprising an expression control sequence operatively linked to a nucleic acid of interest, wherein said expression control sequence is heterologous to and governs seed-specific expression of said nucleic acid of interest, and wherein said expression control sequence is at least 95% identical to the nucleic acid sequence SEQ ID NO: 22; wherein said expression control sequence governs seed-specific expression of said operatively linked nucleic acid of interest.
 2. The expression cassette of claim 1, wherein said expression control sequence comprises the nucleic acid sequence SEQ ID NO:
 22. 3. A vector comprising the expression cassette of claim
 1. 4. The vector of claim 3, wherein said vector is an expression vector.
 5. A non-human host cell comprising the expression cassette of claim 1 or a vector comprising said expression cassette.
 6. A non-human transgenic organism comprising the expression cassette of claim 1 or a vector comprising said polynucleotide.
 7. A method of expressing a nucleic acid of interest in a host cell comprising: (a) introducing an expression cassette comprising an expression control sequence operatively inked to a nucleic acid of interest, wherein said expression control sequence is heterologous to and governs seed-specific expression of said nucleic acid of interest, and wherein said expression control sequence is at least 95% identical to the nucleic acid sequence of SEQ ID NO: 22 or 25 or a vector comprising said expression control sequence, into a host cell; and (b) expressing said nucleic acid of interest in said host cell.
 8. The method of claim 7, wherein said host cell is a plant cell.
 9. A method of expressing a nucleic acid of interest in a non-human organism comprising: (a) introducing an expression cassette comprising an expression control sequence operatively inked to a nucleic acid of interest, wherein said expression control sequence is heterologous to and governs seed-specific expression of said nucleic acid of interest, and wherein said expression control sequence is at least 95% identical to the nucleic acid sequence of SEQ ID NO: 22 or 25 or a vector comprising said expression control sequence, into a non-human organism; and (b) expressing said nucleic acid of interest in said non-human transgenic organism.
 10. The method of claim 9, wherein said non-human transgenic organism is a plant or seed thereof.
 11. The method of claim 9, wherein said nucleic acid of interest is expressed seed-specifically.
 12. The method of claim 7, wherein said nucleic acid of interest is expressed seed-specifically.
 13. The expression cassette of claim 1, wherein said expression control sequence is at least 98% identical to the nucleic acid sequence SEQ ID NO:
 22. 14. The expression cassette of claim 1, wherein said expression control sequence is at least 99% identical to the nucleic acid sequence SEQ ID NO:
 22. 